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(HD) Never Can Say Goodbye - Jackson 5 Multitrack Cover (videosong) - jibcraip MP3
http://tinyurl.com/entertaineramazon http://tinyurl.com/iTunesentertainermp3 Hey All !! Here's a new video !! No a capella stuff this time...This is one of my favorite ...
The Jackson 5 - Never Can Say Goodbye (Mono Mix) (Slayd5000) MP3
Slayd5000 presents... The Jackson 5 - Never Can Say Goodbye (Mono Mix) (Promo 45) N-joi! & subscribe to my channel. Positive comments are always ...
Jackson Five-I Never Can Say Goodbye MP3
Clip from The Jacksons an american dream.
The Jackson 5 - Never Can Say Goodbye (A Cappella) [Audio HQ] HD MP3
The Jackson 5 - Never Can Say Goodbye (A Cappella) Source: EP "Pure Michael: Motown A Cappella" Motown 2009 Motown is the owner of the audio, not me.
The Jackson 5-Never can say goodbye MP3
R.I.P Michael Jackson Jackson 5 Chorus: Never can say goodbye No no no no, I Never can say goodbye Even though the pain and heartache Seems to follow ...
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Tema de Jackson5 cantada por la dulce voz de Michael Jackson... Una linda letra que te robará el corazón....
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Flip Wilson Show (4.11.1971)
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Never Can Say Goodbye in the style of The Jackson 5 karaoke with karaoke video lyrics MP3
Download "Never Can Say Goodbye" in the style of The Jackson 5 in MP4 or MP3+G formats available here: https://karaoke.stingray.com/search/song?
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E-Nyce explains how he sampled Jackson 5 Never can say goodbye.
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My name is Vachelle Gil and this is a tribute to my hero, Michael Jackson, rest in peace. The song is called Never Can Say Goodbye by The Jackson 5. I hope ...
Jackson 5 - Never Can Say Goodbye MP3
One of my favorite songs of Jackson 5.This is from the Jackson 5 fouth studio album Maybe Tomorrow.Enjoy the video.
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Who doesn't love the Jackson 5? More importantly who doesn't love Motown? Like What You See? Follow Me on Twitter: @MatthewTParsons Follow Me on ...
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A mashup song, created by DJ Zebra, of the Jackson 5's "Never Can Say Goodbye" Lenny Kravitz's "It Ain't Over Till It's Over" and Marvin Gaye's "Let's Get It On." ...
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Inspired by Stevie Wonder's live version. Just wanted to practice some talkbox, so I made this cover of some old jams. Close To You (lots of people) and Never ...
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The Miller Brothers.
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Released in 1993 by Motown Records. *Any Negative Comments Will Get You Blocked & The Comment Removed*
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Never Can Say Goodbye (Clifton Davis / Jackson Five / Grant Green) Hammond Grooves: Daniel Latorre: Hammond B-3 e caixa Leslie Wagner Vasconcelos: ...
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The Jackson 5- Never Can Say Goodbye MP3
The Jackson 5 on the Flip Wilson show singing Never Can Say Goodbye. This is a nice remix they did on the show, and the dancing is awesome. Thanks cdog.


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Like any braised dish, this lamb shanks recipe is better when made one day ahead.
Where most chefs used to just reach for kosher salt, there are now many paths to savory goodness.
We've flavored the lamb shanks with North African spices and woken up the rich meat and vegetables with a handful of pomegranate seeds and fresh mint leaves sprinkled on just before serving.
This lamb shank recipe calls for cooking them uncovered in their broth. It simultaneously browns and braises them, adding richness and color. Braise them a day ahead; the flavor will deepen overnight in the fridge.
Add the vegetables partway through braising to ensure they will have enough time to be infused with meaty flavor without overcooking. Serve with a loaf of your favorite crusty bread for sopping up the juices.
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With writer/director Adam Gierasch's noir horror thriller Schism set for its world premiere Saturday, October 12, at 7:30pm at Screamfest LA, read on for our chat with the filmmaker and star Ashlynn Yennie as well as a look at some stills!
Produced by Gierasch, Jace Anderson, Kate Hoffman and Ray Markovich, Schism stars Yennie (The Human Centipede), Callum Blue (Columbiana), Vinnie Jones (The Midnight Meat Train) and Nicole LaLiberte (Girls Against Boys) and revolves around Dylan White (Blue), a New Orleans native who leads a normal, boring life until horrific visions begin to interrupt his waking moments and he's forced to face the fact that he literally doesn't know who he is.
Chatting with Schism director Gierasch, who along with his wife and long-time writing partner, Jace Anderson, penned the flick (the duo's previous writing credits include the horror features The Mother of Tears, Autopsy and the Night of the Demons redux, the latter two he helmed), the filmmaker stated of this most recent foray into bad men, bad dames, bad sex and bad intentions, "Schism is my most personal film so far, and I've been wanting to make it forever. I didn't think I'd ever get the chance."
Of the film's inception, "It all started in the Burbank airport," he said. "Jace and I were going to somewhere in Texas to visit her family for Thanksgiving, and it was one of those awful trips involving layovers and plane changes; pure hell. I really didn't want to go. Basically, I pressured her into not getting on the plane, and we stayed in LA for the weekend. On the way home all I could think was, 'Am I damned for doing this?' That's where the idea came from. I started thinking about stories involving guilt and redemption. Can you change your fate, or are you damned forever once you fuck up? Can you ever truly make amends? I should mention that I'm a nice Jewish boy from upstate New York, so while the guilt part makes sense, the Christian overtones don't."
"As for the writing process," Gierasch continued, "it was the same as it always is. I do structure and story, and Jace puts it on the page. I will say that in the earlier drafts some of the set pieces were even more graphic than they are now, and Jace refused to put the phrase 'vaginal lips' in the script. We argued about that one a lot."
Pertaining to the shooting locale of New Orleans and how it informed the tone of Schism, "It's just such a great location," said Gierasch, who had returned to the city for another filmic go (he'd shot Night of the Demons there previously). "There's so much texture and history," he continued. "And there's so much true scuzziness, which was perfect for Schism. The motel where we shot the Dylan and Marlena scenes was truly awful. During the [location] scout we walked into one of the rooms, and there was fresh blood on the bed."
In regards to the casting of actress Ashlynn Yennie (known to horror fans for her turns in the first two The Human Centipede films) in Schism, "I loved her in The Human Centipede and called her in for an audition," offered Gierasch, "and she just lit up the room, which was perfect for the role of Brandy. I can't say enough good things about working with her. She's up for anything and is a joy to have on set."
"Working with Adam and Jace was amazing," actress Yennie chimed in. "I was so honored to be a part of their film, dream and vision for Schism. I originally was asked to audition for the part of Marlena, but they later asked me to read for Brandy, and it was exactly the role I had been looking for. Having done shocking films (The Human Centipede 1 & 2), I wanted to do a part that was more like who I am as a person. Brandy is warm, inviting, caring, sarcastic and strong, and did I mention I got to play opposite the very talented Callum Blue? What girl wouldn't love that?!"
In addition to the Screamfest premiere of Schism, the Richard Greenwood-directed short film The Wretched Prologue, in which Yennie stars, is also slated for its premiere at the fest.
"The Wretched Prologue is awesome," commented the actress, "I mean, who doesn't love a zombie apocalypse with Roman soldiers? I play Claudia, the wife of Marcus, the leader of the Roman Army. Everyone thinks a plague has spread but it's actually zombies, and it's pretty amazing!"
As for the premiere of Schism and his thoughts on the finished film, filmmaker Gierasch stated, "I'm thrilled for it to be in Screamfest and to be premiering here in LA, [but] I'm a neurotic director. One minute I love the film; the next I think it's a disaster. Seriously, I made a weird noir horror art film. And it's pretty terrifying when it's about to come out because of course you want everyone to love it, but the intent was always for it to be this edgy, weird, cross-genre thing. So hopefully that's what it is."
For more on Screamfest, which runs October 8th to the 17th at the Laemmle NoHo 7 (5240 Lankershim Blvd., North Hollywood, CA 91601), visit Screamfest's website, "like" Screamfest on Facebook, and follow Screamfest on Twitter.
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UK Dating
Hi can you please cancel my membership for the time being After a thoroughly enjoyable year meeting up some really interesting people I've met a... more..
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Prince Albert of Monaco and Charlene Wittstock attend lavish wedding receptionPosted on: July 8, 2011 11:44:29
Lonely hearts searching for a new partner using online dating websites could be interested in the latest developments concerning Prince Albert of Monaco and Charlene Wittstock.
The couple, who were married in a lavish ceremony last week, looked happier than ever as they attended their wedding reception in South Africa yesterday (July 7th).
Despite rumours the nuptials may not go ahead, the couple enjoyed a passionate kiss in front of the cameras at the swanky Oyster Box Hotel in Durban.
Stunning Wittstock had reportedly made several attempts to run away prior to the ceremony after hearing her fiance had fathered a third love child.
The former Olympic swimmer allegedly had her passport confiscated by Monaco police at the airport in a bid to prevent her from leaving the country.
Prince Albert is well-known for his love of beautiful women and has previously been romantically linked to supermodel Claudia Schiffer and actress Broke Shields.
Looking to find men? Try elite dating!
Related Online Dating News
Hong Kong Phil boss "dreams of working with Jackie Chan"Posted on: April 17, 2014 16:26:26
The musical director of one of Asia rsquo s most well-known orchestras dreams of bringing in some unorthodox talent to perform with his musicians Speaking to the South China Morning...
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Lee Ryan and Casey Batchelor steam up the CBB housePosted on: January 6, 2014 17:25:01
They may have been shackled together when they entered the house but Lee Ryan and Casey Batchelor have done anything but drift apart since they were set free In fact...
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The online dating racePosted on: January 6, 2014 16:22:05
New research has revealed how likely single people are to get a message from an online dating site throughout According to MailOnline age matters more to men than women as...
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When Danielle Cornwall-Nash appeared on ITV s prime time dating show Take Me Out last year she hoped that she would find a new partner and maybe even a new...
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Welcome dinner with the hostess of Global Gift, Eva Longoria and María Bravo @ Finca Besaya, Marbella, July 2017, Gallery vol. 2
Starlight Events is proud to present the Gladiator World Cup 2012. The very first time that a K1 tournament takes place in a bullring.
Event infoHost: Starlight Events
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Pseudo attP sites in favor of transgene integration and expression in cultured porcine cells identified by streptomyces phage phiC31 integrase
DNA replication and genome stability
Phage PhiC31 integrase integrates attB-containing plasmid into pseudo attP site in eukaryotic genomes in a unidirectional site-specific manner and maintains robust transgene expression. Few studies, however, explore its potential in livestock. This study aims to discover the molecular basis of PhiC31 integrase-mediated site-specific recombination in pig cells. We show that PhiC31 integrase can mediate site-specific transgene integration into the genome of pig kidney PK15 cells. Intramolecular recombination in pig PK15 cell line occurred at maximum frequency of 82% with transiently transfected attB- and attP-containing plasmids. An optimal molar ratio of pCMV-Int to pEGFP-N1-attB at 5:1 was observed for maximum number of cell clones under drug selection. Four candidate pseudo attP sites were identified by TAIL-PCR from those cell clones with single-copy transgene integration. Two of them gave rise to higher integration frequency occurred at 33%. 5′ and 3′ junction PCR showed that transgene integration mediated by PhiC31 integrase was mono-allelic. Micro- deletion and insertion were observed by sequencing the integration border, indicating that double strand break was induced by the recombination. We then constructed rescue reporter plasmids by ABI-REC cloning of the four pseudo attP sites into pBCPB + plasmid. Transfection of these rescue plasmids and pCMV-Int resulted in expected intramolecular recombination between attB and pseudo attP sites. This proved that the endogenous pseudo attP sites were functional substrates for PhiC31 integrase-mediated site-specific recombination. Two pseudo attP sites maintained robust extracellular and intracellular EGFP expression. Alamar blue assay showed that transgene integration into these specific sites had little effect on cell proliferation. This is the first report to document the potential use of PhiC31 integrase to mediate site-specific recombination in pig cells. Our work established an ideal model to study the position effect of identical transgene located in diverse chromosomal contexts. These findings also form the basis for targeted pig genome engineering and may be used to produce genetically modified pigs for agricultural and biomedical uses.
Transgene technology holds promising applications in biomedicine and agriculture. Classical DNA pronuclear microinjection is a reliable tool to produce transgenic animals, but the inefficiency and uncontrollability of integration site and copy number are the major limitations
These problems can be overcome by targeting the transgene to a specific genomic site via homologous recombination (HR). However, it is usually labor-intensive and time-consuming due to the extremely low frequency of HR in mammalian cells
Phage integrases carry out irreversible and unidirectional recombination between attachment sites of phage and bacteria genomes, known as attB and attP sites, respectively
An optimal molar ratio of PhiC31 integrase to substrate is important to its activity in pig cell environment
To know if PhiC31 integrase system functions in pig cell environment, we used pCMV-Int and pBCPB+ plasmids to detect extra chromosomal intramolecular recombination. The two plasmids contain the required components of PhiC31 system: the integrase, attB and attP recognition substrates placed in direct orientation. In this context, PhiC31 integrase will catalyze the recombination between attP and attB site, deleting LacZ gene and yielding attL and attR hybrid sites. Forty-eight hours post transfection of the two plasmids into pig kidney PK15 cell line, a cell PCR assay was conducted to detect attL and attR sites with specific primers. Expected products were only obtained from the co-transfection of both the two plasmids. Neither pCMV-Int nor pBCPB+ produced these amplicons. We also titrated the molar ratio between enzyme and substrate. It showed that molar ratio of 5:1 (pCMV-Int: pBCPB+ = 5:1) resulted in maximum recombination efficiency 82%. The result suggested that the three components of PhiC31 integrase system were sufficient to induce site-specific intramolecular recombination in pig cells, implying that no cofactors were required for PhiC31 integrase to function in pig cells.
To test if PhiC31 integrase is capable of catalyzing intermolecular recombination, we constructed pEGFP-N1-attB (Figure
Figure 1
PhiC31 integrase is active in pig cells
PhiC31 integrase is active in pig cells. (A) Mechanism of action of PhiC31 integrase-mediated site-specific recombination. The integrase catalyzes the intramolecular recombination between attB and attP sites within a single plasmid, producing two mini-circle plasmids with hybrid attR and attL sites, respectively. (B) PhiC31 integrase is functional in pig cells. Transfection of pig PK15 cells were performed with three different ratios of PhiC31 integrase plasmid to reporter pBCPB+. Resultant attL and attR sites were examined by PCR. The indicated recombination efficiency was calculated by measuring signal intensity in Visioncapture with the following formula: recombination efficiency = Intensity of attL/Intensity of (attL and pBCPB + backbone). A 5:1 ratio had 82% recombination efficiency. (C) The map of pEGFP-N1-attB (5138 bp). A 400 bp attB sequence was sub-cloned into the AseI site of pEGFP-N1, resulting in the site-specific integration reporter pEGFP-N1-attB. (D) EGFP expression level in a time lapse. The y-axis shows the expression levels and x-axis shows days after transfection. Four groups of plasmids (pEGFP-N1, pEGFP-N1-attB, pEGFP-N1/PhiC31 and pEGFP-N1-attB/PhiC31) were transfected into PK15 cells. EGFP expression level was quantified by measuring the extracellular EGFP by a Glomax Multi Jr fluorescence detector. IGF-I was detected by ELISA and used as an internal control for data normalization. pEGFP-N1-attB/PhiC31 maintained high EGFP level in comparison to other settings. (E) An optimal molar ratio of PhiC31 and pEGFP-C1-attB plasmids produced more cell clones expressing EGFP. Various ratios of PhiC31 and pEGFP-N1-attB plasmids were transfected into PK15 cells and cell clones were selected by G418. The y-axis shows % of green to total clones and x-axis shows different molar ratios. A 5:1 ratio produced more cell clones expressing EGFP, significant higher than other settings, including random integration (0/1 ratio). *, p < 0.01.
Identification of candidate pig pseudo attP sites
Next, we sought to identify the integration sites of reporter plasmid in the genome of PK15 cells. First, we used quantitative real-time PCR to screen single-copy transgene integrated cell clones. This manipulation not only allowed us to identify the precise location of candidate pseudo attP site but also to compare characteristics of the integration sites. Total number of 34 cell clones was picked from the transfection groups of different molar ratios. From these, eleven single-copy transgene integrated cell clones were identified, where the single-copy transgene integration frequency among G418-resistant cell clones is 33%. TAIL-PCR was then performed to isolate the integration sites
Figure 2
Isolation of pig pseudo attP sites
Isolation of pig pseudo attP sites. (A) Screening of transgenic cell lines with single-copy transgene integration by quantitative PCR. 34 transgenic cell lines expressing EGFP (shown in x-axis) were screened for single-copy transgene integration within PK15 cellular genome. This assay demonstrated that 11 cell lines harbored a single-copy pEGFP-N1-attB. The other 23 cell lines had distinct copy numbers of pEGFP-C1-attB, varying from 2 copies to 15 copies. (B) Isolation of integration sites. Left, microscopic images of four transgenic cell clones under fluorescence field (10×). Middle, TAIL-PCR was performed to isolate the integration sites. Right, Specific PCR products were TA cloned, sequenced and aligned by BALT against pig genome (UCSC). Red arrow represents pig genome sequence, and green arrow represents pEGFP-N1-attB sequence. The lanes shown as 1st, 2nd and 3rd represent the first, second and third round TAIL-PCR products fractionized by 1.5% agarose gel. M is 1 kb DNA molecular marker (Fermentas).
In order to prove the integrity of site-specific recombination, junction PCR was carried out to detect the specificity of DNA sequence comprised of pig genome and reporter plasmid. A specific DNA product was detected for both attL and attR sequences of all the four pseudo sites (Figure
Figure 3
Confirmation of integration sites and allele analysis
Confirmation of integration sites and allele analysis. (A) attR and attL detection by junction PCR. Integration site and its neighboring sequences were retrieved from UCSC Pig Genome. Primers were designed across the junction site. PCR analysis revealed that pEGFP-N1-attB had been integrated into the sites identified by TAIL-PCR. In addition, all the 4 integration events occurred to one chromosome as demonstrated, implying that the transgene is integrated as single-allele at the 4 pseudo attP sites. (B) Micro insertion or deletion at integration site. PCR products in Figure
Sequencing of the TAIL-PCR products further revealed the covalent linkage of reporter plasmid in PK15 cellular genome. The joining sites were in accordance with PhiC31 integrase-mediated attB and attP crossover reaction. It implies that PhiC31 integrase functions in pig cells to introduce attB-containing plasmid into pseudo attP sites. Meanwhile, we observed imperfect repairing in the junction sites (Figure
Functional rescue assay proved that candidate pig pseudo attP sites were
After identification of candidate pseudo attP sites in PK15 cellular genome, we then asked if they were functional to reconstitute site-specific recombination in vitro. To this end, we created reporter plasmid by cloning pig pseudo attP sites into pBCPB+ plasmid, replacing wild-type attP sequence. As restriction sites were unavailable besides wild-type attP site, it was achieved by ABI-REC cloning method (Figure
Figure 4
Candidate pig pseudo attP sites can reconstitute site-specific recombination in a functional rescue assay
Candidate pig pseudo attP sites can reconstitute site-specific recombination in a functional rescue assay. (A) Scheme of reporter plasmid construction. Pig pseudo attP sites were cloned into pBCPB+ plasmid to replace the wild-type attP site by ABI-REC. The resultant plasmid was named p'BCPB+. This resulted in 4 p'BCPB+ plasmids, i.e. 5113, 5156, 1015, and 2015, respectively. p'BCPB+ was used to reconstitute the attB-pseudo attP recombination with the presence of PhiC31 integrase. For the ABI-REC details, please refer to additional files. (B) Rescue assay. PhiC31 integrase plasmid, pBCPB+, and 4 p'BCPB+ were transfected into PK15 cells. 48 h post transfection, genomic DNA was extracted and used for PCR detection of attR hybrid site with attR-F and attR-R primers. Pig endogenous MSTN gene (500 bp) was used as internal control to normalize the quantity of genomic DNA template. Primers attL-F and attR-R (555 bp) were used to quantify the amount of pBCPB+. (C) Recombination efficiency. The VisionCapture tool was used to quantify the density of the PCR product in (B). Recombination efficiency was calculated with the following formulation: Amount of attR/(amount of attR + amount of pBCPB+). It shows that the recombination efficiency of wild-type attP and attB sites was up to 80%. Pig 5113 pseudo attP site has a recombination efficiency of 60%, higher than the other 3 pig pseudo attP sites.
Additional file 1: Figure S1
ABI-REC of pig pseudo attP sites into pBCPB+ plasmid. Fused products were indicated by arrow. M is molecular DNA ladder. 5113, 5156, 1015 and 2015 represents four pig pseudo attP sites cloned by ABI-REC.
Click here for file
Additional file 2: Figure S2
p'BCPB+ plasmid restriction analysis. Recombinant plasmids were analyzed by restriction digestion. M is molecular DNA ladder. 5113, 5156, 1015 and 2015 stands for the four recombinant plasmids derived from backbone pBCPB+. For each plasmid, the first lane shows undigested DNA and the second lane shows the restricted DNA.
Click here for file
Additional file 3: Figure S3
Partial sequences of four p'BCPB+ plasmids were shown. Native restriction site is underlined. Cloned pig pseudo attP site is shown in red, where the identified pseudo attP site is shown in bold. pBCPB+ plasmid sequence is shown in black. Recombination crossover is shown in bold and italics.
Click here for file
Pig pseudo attP sites are in favor of robust transgene expression
We evaluated the favorability of the four pseudo attP sites to transgene expression. Extracellular EGFP expression was measured by a fluorescence detector. IGF-I was used as internal control to normalize the input variation as IGF-I was secreted into medium. It demonstrated that 5113 and 2015 pseudo attP sites were in favor of robust EGFP expression, significantly higher than that of 5156 and 1015 sites (Figure
Figure 5
Pig pseudo attP sites are in favor of robust transgene expression
Pig pseudo attP sites are in favor of robust transgene expression. (A) Extracellular EGFP expression. Medium containing extracellular EGFP was harvested for EGFP fluorescence and IGF-I detection. IGF-I was internal control for input normalization. IGF-I concentration was measured by a porcine ELISA kit. Sp, the supernatant of PK15 cell line as negative control. (B) Intracellular EGFP level by western blot. Transgenic cells were harvested and quantified by a BCA protein quantification kit. Anti-EGFP antibody was used to detect the intracellular EGFP expression, and anti-β-actin antibody was used as internal control. MW of EGFP and pig β-actin is 27 kD and 43 kD, respectively. They are shown by arrows. PK15 cell lysate was used as negative control. The intensity of the bands was quantified by VisionCapture and converted into Prism to create the bar graph. (C) Cell proliferation assay. 103 cells were seeded in a 96-well plate in triplicate. 10 μl Alamarblue indicator (Gibco) was added into the medium (with a final concentration of 10% v/v). The plate was incubated for additional 5 hours. The optical densities were measured at 570 nm and 630 nm in a micro-plate reader. Reduction percentage was calculated according to the formula provided in the instruction. Such a procedure was repeated in continuous 15 days. The reduction of four time points (day1, day5, day10 and day15) was indicated. Fresh medium was used as blank control. PK15 is the wild-type cell line. R1, R2 and R3 are randomly integrated transgenic cell lines.
In this paper we demonstrated the catalyzing activity of PhiC31 integrase to introduce site-specific transgene integration into PK15 cellular genome. We showed that PhiC31 integrase system work synergistically in a quantitative manner. For the extra chromosomal site-specific recombination, it was found that the optimal ratio of pCMV-Int (6230bp) to pBCPB+ (7221bp) is 5:1 in pig cellular environment. PhiC31 integrase catalyzes the recombination as high as 82%. In an attempt to examine the potency of PhiC31 integrase-mediated intermolecular recombination, we also observed that a 5:1 molar ratio of pCMV-Int to pEGFP-N1-attB resulted in robust and long term EGFP expression and produced maximum percentage of EGFP-positive cell clones. Please note that in all the transfections we have assured the charge ratio of DNA plasmid to transfection reagent invariable. Hence, our results allow for a direct comparison among diverse transfection settings. Such results were also observed in previous reports
Secondly, as documented previously in human, mouse, rat, fruit fly, silkworm and cattle, pseudo attP site profile varies across species
Thirdly, we reconstituted the site-specific recombination by performing a functional rescue assay. To our knowledge, this is the first report to prove candidate pseudo attP site to be
Finally, we carried out TAIL-PCR to isolate the integration site. In previous studies, inverse PCR, half-nested PCR and plasmid rescue were often used to identify the candidate pseudo attP sites. But the common drawback of these methods is that they require a large starting amount of genomic DNA (usually 10 μg) for restriction and ligation. However, TAIL-PCR needs as little as 0.5 μg genomic DNA. In this work, we obtained four pig pseudo attP sites by TAIL-PCR. Moreover, the integration of reporter plasmid into all these four pseudo attP sites could be proved by junction PCR, indicating that the positive rate of TAIL-PCR is 100%. Accordingly, we believe that TAIL-PCR is a valuable tool owing to its ease, low cost and high reproducibility.
Our data demonstrated that PhiC31 integrase functions efficiently for site-specific transgene integration into PK15 cellular genome. We isolated four pig pseudo attP sites and proved their recombination activity by a functional rescue assay. PhiC31 integrase induces mono-allelic transgene integration in pig cells. Pig pseudo attP sites were proved to be in favor of robust transgene expression. Our findings also established an ideal model to study the position effect of identical transgene structure located in diverse chromosomal contexts. These results form the basis for targeted pig genome engineering and may be used to produce genetically modified pigs for agricultural and biomedical uses.
Plasmid construction and DNA manipulation
The plasmid pCMV–Int (expressing PhiC31 integrase) and reporter plasmid pBCPB+ were gifts from M. P. Calos (Stanford University, USA). A Roche high pure PCR template preparation kit was used to extract genomic DNA. All plasmids were prepared by a TIANpure midi plasmid kit (Tiangen, Beijing, China). A Nanophotometer P-class was used to check the quality and quantity of DNA (Implen, Germany). AttB fragment was amplified from pBCPB+ and inserted into the AseI site of pEGFP-N1 (Clontech), and the resultant plasmid was pEGFP-N1-attB. For ABI-REC to create p'BCPB+ plasmids, pig pseudo attP sites were amplified by a KOD plus high-fidelity DNA polymerase and fused into pBCPB+ plasmid, replacing the wild-type attP site. For details of ABI-REC protocol, please refer to previous description
Cell culture, transfection and stable cell line selection
Pig kidney PK15 cell line was cultured in DMEM (Dulbecco's modified Eagle's medium, Gibco, Life Technologies) supplemented with 10% (v/v) FCS (fetal calf serum, Hyclone), 100 IU/ml penicillin and 0.1 mg/ml streptomycin. Cells were grown at 39°C in the presence of 5% CO2 and were subcultured (1:5) every 3 days. To obtain stable transfected cells, PK15 cells were split in a 35-mm plate until they reached 80% confluency. Cells were washed with PBS and then incubated with a mixture of 4 μg of DNA, 10 μl of Lipofectamine 2000 (Invitrogen) and 500 μl of Opti-MEM that was prepared as described by the manufacturer. To ensure the constant charge ratio of total nucleic acid and lipid, "filler or unrelated DNA" (pUC19 plasmid) was added to the transfection mixture. Following 6 h incubation, the medium was replaced with fresh medium. 48h post transfection, 600 μg/ml G418 (Sigma) was added. After 2 weeks of selection, G418-resistant colonies were picked manually and maintained in media with 50 μg/ml of G418. For cell proliferation assay, an Alamar blue kit (Gibco) was used. Briefly, 1000 cells were seeded in a 96-well plate in triplicate. 10 μl Alamar blue indicator was added into the medium (with a final concentration of 10% v/v). The plate was incubated for additional 5 hours. The optical densities were measured at 570 nm and 630 nm in a micro-plate reader (Multiskan MS; Labsystems, Helsinki, Finland). Reduction percentage was calculated according to the formula provided in the instruction (Gibco). Fresh medium was used as blank control. Such a procedure was repeatedly performed for continuous 15 days. The reduction values at day1, day5, day10 and day 15 were shown in the manuscript (Figure
Extracellular EGFP quantification
Extracellular EGFP into medium was quantified using a Glomax Multi Jr fluorescence detector, according to the manufacture's manual book. The EGFP level was indicated in arbitrary units. In order to normalize the input variation, an ELISA assay was used to measure the IGF-I expression level secreted into the medium as an internal control. Briefly, a 96-well plate was coated with anti-pig IGF-I antibody (Abcam, Cambridge, UK). After blocking, extracellular proteins were added in triplicate. IGF-I expression was then determined using a horseradish peroxidase-conjugated secondary antibody (Abcam) with 3, 3′, 5, 5′-tetramethylbenzidine (Sigma-Aldrich) as the substrate. The reaction was stopped by addition of 100 μL of 1 M H2SO4. Absorbance was measured at 450 nm using an ELISA reader (Bio-TEK Instruments Inc, Winooski, VT, USA).
Western blot
The cells were washed thrice with ice-cold PBS to remove residual medium as much as possible. Afterwards the cells were lysed by NP40 Cell Lysis Buffer (Invitrogen) with phenylmethylsulfonyl fluoride (PMSF). After centrifugation, the supernatants were collected and quantified by Pierce BCA protein assay kit (Thermo Scientific). The total proteins were denatured by boiling for 5 minutes in a water bath and chilled in ice. 15% SDS-PAGE was used to size-fractionate the samples. Then they were transferred electrophoretically onto polyvinylidene difluoride membranes (Amersham Biosciences, Piscataway, NJ). After blocking with 2% milk, the membranes were incubated with primary antibodies at 4°C for 1 hour. The monoclonal primary antibody against EGFP (1:1000) was purchased from ProteinTech Co. Ltd. (Wuhan, China). The monoclonal primary antibody against β-actin (1:10000) was purchased from Sigma. After washing with PBST, the membranes were incubated with rabbit anti-mouse horseradish peroxidase-conjugated secondary antibodies (Sigma) for 2 h at room temperature. The signals were visualized by enhanced chemiluminescence (Perkin-Elmer, Norwalk, CT).
Genome walking
Genome walking was carried out by a TAIL-PCR kit (Takara) according to the manufacture's instruction. Briefly, 0.5 μg PK15 genomic DNA was used to perform the first round PCR with AP1, AP2, AP3, AP4 and IV-R1 primers in a 50 μl reaction, respectively. 1μl PCR products were used as template for 2nd and 3rd round PCR with IV-R2 and IV-R3 primers to obtain specific PCR bands. The DNA bands were excised and TA cloned into T vectors (Takara). Sanger sequencing was conducted in a ABI3730 DNA Analyzer (Applied Biosystems, Foster City, CA, U.S.) and the sequences were assembled and analyzed using DNAStar software (DNASTAR Inc., Madison, WI. U.S.). All primers are listed in Table
Real-time PCR
A Toyobo real-time SYBR Green I PCR master mix was used to amplify EGFP or TFRC gene in a Rotor Gene 6000 real-time rotary analyzer (Corbette Lifescience, Australia). The 20 μl PCR reaction mixture contained 10 μl 2 × master mix, 0.3 μl primer mix (5 μM each), 0.5 μl genomic DNA (50 ng) and 9.2 μl PCR-grade water. A two-step amplification protocol was used with the following parameters: 95°C for 2 minutes to pre-denature the template and activate Taq DNA polymerase followed by 40 cycles each of denaturation at 95°C for 8 seconds, annealing and extension at 60°C for 25 seconds. A final melting temperature analysis from 50°C to 99°C was used to ensure amplicon uniformity. Fluorescence was acquired at the step of annealing and extension. All PCR amplifications were performed in triplicate for each treatment. Wild-type PK15 genomic DNA was used as negative control. EGFP standard curve was produced by amplifying pEGFP-N1-attB at gradient concentration of 10 ng, 1 ng, 100 pg, 10 pg, 1 pg. TFRC standard curve was produced by amplifying wild-type gDNA at gradient concentration of 100 ng, 10 ng, 1 ng, 100 pg, 10 pg. Primers used for real-time PCR (Table
Statistical and bioinformatic analysis
The results of colony calculation, real-time PCR, EGFP expression were shown as the mean ± SD (n = 3). The statistical analysis was performed using
TAIL-PCR: Thermal asymmetric interlaced PCR; ABI-REC: Asymmetric bridge PCR and intramolecular homologous recombination; EGFP: Enhanced green fluorescence protein; PBS: Phosphate-buffered saline; attB: Attachment site of bacteria; attP: Attachment site of phage; attL: Left attachment site; attR: Right attachment site; ELISA: Enzyme-linked immunosorbent assay; IGF-I: Insulin-like growth factor I; TFRC: Transferrin receptor.
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
YZ conceived the project, designed all the primers, analyzed all the experimental data and wrote the manuscript. XL performed cell culture, transfection and selection. LZ carried out TAIL-PCR. CS conducted immunoblotting. ZM performed ELISA assay. ZH performed TA cloning and sequence analysis. LZ performed PCR and real-time PCR. LL and WH were involved in plasmid preparation and keeping stocks. HX and QW performed cell proliferation assay. XZ proposed the functional rescue of pig pseudo attP site by ABI-REC method. All authors read and approved the final manuscript.
The authors are grateful to M. P. Calos (Stanford University, USA) for her generosity to provide us expression plasmids. We are also grateful to Dr. Qingwen Ma (Shanghai Jiaotong University) and Dr. Ruhong Jiang (Applied Stem Cells, Inc., USA) for their helpful discussions. We would like to thank the assistance of team of veterinary medicine from Hubei Academy of Agricultural Science in ELISA assay. This study was supported by Natural Science Foundation of China (31201790), International Science and Technology Cooperation Project of Hubei Province (2009BFA012), a Key Project of Youth Foundation (2011NKYJJ13) of Hubei Academy of Agricultural Science (HAAS), Innovation Center for Agricultural Sciences and Technologies of Hubei Province (2011-620-001-003), open grant from Hubei Key Laboratory of Animal Embryo Engineering and Molecular Breeding (2013ZD120/121). It was also supported by China Major Program of Genetically Modified Organism New Species Cultivation (2013ZX08010-003, 2013ZX08006-003, and 2013ZX08006-002).
Animal transgenesis: an overview
PiggyBac transposon-mediated gene transfer in human cells
Efficient transgenesis in farm animals by lentiviral vectors
Pig transgenesis by Sleeping Beauty DNA transposition
Strategies for selection marker-free swine transgenesis using the Sleeping Beauty transposon system
Germline transgenic pigs by sleeping beauty transposition in porcine zygotes and targeted integration in the Pig genome
Position effect variegation and imprinting of transgenes in lymphocytes
Gene targeting using a promoterless gene trap vector ("targeted trapping") is an efficient method to mutate a large fraction of genes
Targeted integration in rat and mouse embryos with zinc-finger nucleases
Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression
In vitro site-specific integration of bacteriophage DNA catalyzed by a recombinase of the resolvase/invertase family
Construction of transgenic Drosophila by using the site-specific integrase from phage PhiC31
Site-specific gene integration in cultured silkworm cells mediated by PhiC31 integrase
Site-specific genomic integration produces therapeutic Factor IX levels in mice
PhiC31 integrase confers genomic integration and long-term transgene expression in rat retina
Site-specific integrase-mediated transgenesis in mice via pronuclear injection
Identification of pseudo attP sites for phage PhiC31 integrase in bovine genome
A PhiC31 integrase‒mediated integration hotspot in favor of transgene expression exists in the bovine genome
A site-specific recombinase-based method to produce antibiotic selectable marker free transgenic cattle
Efficient isolation and mapping of Arabidopsis thaliana T-DNA insert junctions by thermal asymmetric interlaced PCR
A direct comparison of two nonviral gene therapy vectors for somatic integration: in vivo evaluation of the bacteriophage integrase PhiC31 and the sleeping beauty transposase
An asymmetric PCR-based, reliable and rapid single-tube native DNA engineering strategy
A profile of native integration sites used by PhiC31 integrase in the bovine genome
Transcription start regions in the human genome are favored targets for MLV integration
Precision genetic engineering in large mammals
Precision editing of large animal genomes
ΦC31 integrase interacts with TTRAP and inhibits NFκB activation
Mutational analysis of highly conserved residues in the phage phiC31 integrase reveals key amino acids necessary for the DNA recombination
One-stop polymerase chain reaction (PCR): An improved PCR method with speedy operation and comparable efficiency
Involvement of HFq protein in the post-transcriptional regulation of E. coli bacterial cytoskeleton and cell division proteins


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Now in our 51st year, the Georgia Farm Monitor is the only national and state weekly news and information television program dedicated to Georgia's largest industry - agriculture. In that time we've produced more than 2,500 weekly episodes!
About The Monitor
The Georgia Farm Monitor is produced by the Georgia Farm Bureau, the state's largest general farm organization. From our offices in Macon, the Farm Monitor staff travels the state, the Southeast and to other parts of the country to cover stories of interest to farmers and consumers in this entertaining and informative 30-minute program.
While the program focuses on agriculture with the Georgia and Southeastern farmer in mind, national ag issues, consumer information and interesting feature stories about rural life and people are also part of the show each week.
The show is seen on a network of thirteen stations across Georgia, as well as nationally on the RFD-TV Network.
To find a Farm Monitor station near you, visit this page.
Follow Us!
You can keep up with the latest news and events concerning Georgia agriculture by following the Georgia Farm Monitor on social media. Just click the images below.. we'd love to have you as a follower!
Monitor History
The Monitor has been on the air since 1966, when it began as a joint production of Georgia Farm Bureau and WMAZ-TV in Macon. As the show and staff grew, it eventually moved to its own production studios in the Georgia Farm Bureau state headquarters in 1978 and began adding other TV stations.
Starting in 2000, the Georgia Farm Monitor was one of the first programs to begin airing nationally on the RFD-TV Network, which originates in Nashville, Tennessee. It now airs twice each week on RFD-TV.
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Closed Captioning Certification


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Rakhi Thali with Bounty Chocolates Treat
You will love this Raksha Bandhan combo gift pack for containing everything special that you need to surprise your darling brother on the auspicious occasion. Your brother is always being with you as your best friend,looking out for you and making sure that your are on a safe and correct path of which you are traveling. Rakhis are embossed with colored beads. Also,It comes with a special treat of bounty chocolates. Product Includes : Exclusive Raksha Bandhan Thali,Attractive Rakhi,Bounty Chocolates (Weight : 170 Gms) and Kumkum Chawal in a container for Tikka.


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You've got a slew of footage of family gatherings or dance recitals. Edit those clips into something you can put in a DVD or show at the next reunion. Follow our quick tips to show you the way.
This story, "How to Spruce Up Your Home Videos" was originally published by PCWorld.


text:
2006-07-31 at 2:17:00 pm #16009
Xerox Second-Quarter Net Income Falls 39%; Revenue Gains 1.4%
06– Xerox Corp., the fourth-largest seller of printers and copiers in
the U.S., said second-quarter profit dropped 39 percent.
income fell to $260 million, or 26 cents a share, from $423 million, or
40 cents, a year earlier, when a tax gain buoyed results. Sales rose
1.4 percent to $3.98 billion, the Stamford, Connecticut-based company
said today in a regulatory filing.Chief Executive Officer Anne Mulcahy
is focusing on digital copiers and printers to revive growth and is
seeking to shore up sales of supplies such as toner and paper, which
are more profitable than the machines. She has eliminated 20,000 jobs
since taking over in 2001, paid down $11.5 billion in debt and hired
other companies to make Xerox products."They're getting the installed
base in shape, but they have to get it reflected in earnings per
share," said Shannon Cross, an analyst at Cross Research in Short
Hills, New Jersey. She rates the shares "buy" and said she doesn't
own them. "The challenge is to see growth in post-sale revenue."Cross
expected profit of 24 cents a share, a penny ahead of the 23-cent
average estimate of nine analysts in a survey by Thomson Financial.
Thomson doesn't disclose the basis of its estimates to Bloomberg News.
Xerox in April forecast 22 cents to 24 cents.Xerox shares rose 35 cents
yesterday to $13.65 in New York Stock Exchange composite trading and
have dropped 6.8 percent this year. Shares of Palo Alto,
California-based Hewlett-Packard Co., the largest printer maker, have
gained 8.5 percent in 2006.
Sliding Share
Xerox has fallen to
fourth in both U.S. copier and printer sales, according to Stamford,
Connecticut-based researcher Gartner Inc. The company trails Tokyo's
Canon Inc., Hewlett- Packard and Konica Minolta Holdings Inc. in
copiers and lags behind Hewlett-Packard, Konica Minolta and Round Rock,
Texas- based Dell Inc. in printers.Mulcahy, 53, is shifting to digital
color technology as sales of its analog machines decline. Color pages
are five times as profitable as black-and-white, she said in
April.Almost three-quarters of Xerox's revenue comes from supplies and
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color copiers and printers, systems that run 41 pages per minute,
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Mulcahy's Efforts
Since Mulcahy took over in 2001,
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year was the highest since 1997. In June, Xerox agreed to buy closely
held Amici LLC for $174 million to add research tools for lawyers. It
marked Mulcahy's first acquisition since becoming CEO.She has
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Singapore-based Flextronics International Ltd. and Fuji Xerox Co., a
joint venture with Fuji Photo Film Co. of Tokyo. She also shifted
financial management of equipment leases to Fairfield,
Connecticut-based General Electric Co.


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Although the battle cry has waned slightly and the confetti now only dusts the streets, credit unions wonder... did Bank Transfer Day have a strong impact on reshaping consumer behavior?
Javelin Strategy & Research reports that although over 600,000 consumers have switched from big banks to credit unions, the momentum may be coming to a screeching halt.
Javelin founder, Jim Van Dyke said, "It was a meaningful movement of people from big banks into small community banks and credit unions, but it wasn't a huge number by any stretch."
He adds, "It was quite a feat. I don't think we'll see it repeated any time soon." Why? Van Dyke refers back to the old notion that it takes more than a simple incident to push people to leave their bank and that having to "undo" services like direct deposit and bill pay leave many consumers with that feeling of "why bother."
"Individuals are really resistant to moving their money out of banks," he said. "The difference between what they say and what they do is really different." Javelin tracked and researched consumer banking behavior during a survey in 2003.
He backs this statement by noting that an additional 200,000 consumers stated that they moved their money from one of the top big banks to a credit union due to Bank Transfer Day but ended up taking no action.
Misconceptions and Banker's Opinion
Although moving money can be challenging, misconceptions within the bank-heavy industry continue to rage. Despite the wide availability of member assistance and revolutionary, cutting edge technology offered at most credit unions, Van Dyke claims that young adults are still hesitant leave their big bank because they are under the impression that community banks and credit unions do not offer the same line of "robust online banking tools."
"It turned out that technology and individuals' inertia really won the day in favor of the large banks," Van Dyke says. "If this continues, these small institutions will go the way of the Oldsmobile."
Multi national giants are also not feeling much pressure. American Bankers Association spokeswoman Carol Kaplan says that movement from Bank Transfer Day was miniscule, but made banks sit up and take note.
She says, "While these 600,000 adults represent an exceedingly tiny fragment of the hundreds of millions of Americans with bank accounts, the industry takes consumer sentiment seriously... Consumers have a wealth of choice and banks have always been in favor of competition, regardless of where it comes from."
Although much of the fervor stemmed from Bank of America's $5 debit card fee, Van Dyke says that the giant didn't take a huge hit and losses were similar to what other big banks experienced during Bank Transfer Day. "We didn't see any individual bank punished out of this more than the rest," he says.
During a recent conference, Bank of America CEO Brian T. Moynihan said that the bank experienced a 20% decrease in account closings from fourth quarter 2011 versus the same period in 2010. "So I'd say that yes, we had some impact from the $5 debit fee. That's why we made a decision to reverse it."
Ed Mierzwinski, consumer program director for U.S. PIRG believes that Bank Transfer Day did have an impact. Referring to Moynihan's assertion he said, "Bank of America got the message and did take a hit. Bank of America and other big banks now know that unfair practices will and do result in consumers voting with their feet."
Credit Unions Believe Bank Transfer Day Was Significant
Either way Javelin reports that 5.6 million consumers switched to a credit union in 2011 with only 11% of the switch represented from Bank Transfer Day.
Carolyn Jordan, SVP at Neighborhood Credit Union ($275 million, Dallas/Ft. Worth, TX)refutes Van Dyke's comments about the movement slowing down.
"We have seen continued movement from the megabanks in the Dallas area since Bank Transfer Day. While the debit card fee is no longer an issue, many of the new members who have come to us expressed that after hearing of potential new bank fees, this got them thinking about and looking at credit unions."
As a result Jordan says that Neighborhood is seeing tremendous results in new member activity. "In December we opened 481 new checking accounts which is a 66% increase compared to December of 2010. We had an all time high in January 2012 opening 471 new checking accounts, which is a 58% increase over January 2011."
Jordan also uses a new member comment to demonstrate how bank customers may be under the misimpression that convenience and superior technology aren't available at credit unions.
"One new member I recently talked to said that she was amazed at the fact that we are just as convenient as a bank. She thought that she would have to trade the convenience of ATMs and robust online banking for personal service but has discovered that this is not the case. She told me that she has the benefit of convenience and personal service with us and she just loves it. She expressed that she would never go back to a bank after experiencing the difference of Neighborhood Credit Union. This made me very proud."
Although Bank Transfer Day founder Kristen Christian was not officially part of the Occupy movement, she struck a chord in the hearts and minds of many who are. Occupy San Francisco activist Brian McKeown felt that although protesting sent a message, taking action would have a more significant impact. Late last year he planted the seed of what will be a new credit union in the Bay Area called People's Reserve Credit Union.
Although the credit union is still in its infancy stages, McKeown and his advisor San Francisco entrepreneur Tim Mayer said that while People's Reserve will look and act like a credit union, members may see differences.
Borrowers seeking loans will have to agree to attend special financial literacy courses concentrating on business and personal finance. Special mentors will also be assigned to commercial borrowers to help their business succeed, which will ultimately motivate the borrower to pay back the loan.
In fact McKeown plans to also be diverse with his loan parameters as he may be offering business loans as low as $1,000. He said that the amount could possibly fund materials to make jewelry or allow a business to purchase a used server to create computer apps.
"I'd rather support 12 taco stands than one Taco Bell," he said.
Mayer says, "But there will be a different consciousness," and that the PRCU board "will not focus on shareholder profits," but will be "concerned with the welfare of those they are servicing."
Additional Resources:


text:
So you've decided to break down and buy a new pair of binoculars. You probably even have a favorite manufacturer in mind that you are considering. Even within a single product line, there are a ton of options when it comes to binoculars. Power, objective, water resistant, waterproof, shockproof, eye cups, straps, the list goes on and on.
The list below will help you select the perfect hunting binoculars for your particular situation and style of hunting.
Depending on where you plan to hunt, you will have different considerations in terms of power and objective. For western hunters, 10x or 12x should be adequate for glassing longer distances. Western hunters also need to balance the need for more power with weight. Higher power binoculars are typically larger and heavier than smaller, lower power models. There is a line to dance in terms of weight and power. Consider how much hiking you will be doing and invest in a good pair of binocular straps that evenly distributes the weight across your shoulders, back, and chest.
Eastern bowhunters typically use an 8x-10x power with a large objective (40mm+). In low light, the lower power actually helps you because you have more field of view. Ever spotted something with your naked eye then had trouble finding it when you look through your binos? Some of this may have to do with the power you are using. Lower power binoculars are excellent for poor light and thick woods. Consider your needs and where you will be using your binoculars–sometimes a lower power is the correct power.
When you begin to look online or at outdoor stores for a new pair of binoculars, pay close attention to the following items:
- Warranty
- Lens Manufacturer
- Power & Objective
- Weight
- Eyecup Construction
- Waterproof/Fogproof/Shockproof
From the list, one of the most important items is the warranty program. Customer service and having a great warranty can go a long way to ease your mind in terms of cost and reliability. Some companies provide "no questions asked" replacement of lenses, eyecups, and straps while others require you to jump through a little more red tape. Study each company and determine which has the best warranty program for you.
Why is it so hard to tell in the store?
Similar to buying clothes, don't buy a new pair of binos without "trying them on" first. Aside from online reviews and word of mouth, you will need to actually look through the binos and check for clarity, how much light they gather, how comfortable they are to look through and wear, etc. Everyone's eyes are slightly different and making sure they work with you is key.
It's difficult to differentiate binoculars in the store because most only show their true colors when the light is poor. You need dim light to tell which has better glass and that simply cannot be done standing in a florescent-lit showroom floor.
Tip: Go to the store at twilight or just as it is getting dark. Make friends with the sales person behind the optics counter. Ask, politely, if he or she will accompany you outside so you can really put them to the test. Most stores and employees are happy to help. Going outside when the light is poor will allow you to really test each pair and make a more educated purchase.


text:
|Sarah Charles
|General Information
|Real name:
|Sarah Charles
|First Appearance:
|Tales of the Teen Titans #57 (September, 1985)
|Created by:
| Marv Wolfman
Chuck Patton
Romeo Tanghal
|S.T.A.R. Labs
|Portrayed by:
Dr. Sarah Charles is an intern at S.T.A.R. labs and a University of Michigan college student.
She is the daughter of two very influential parents. Her mother Sasha Charles is a congress-woman in Michigan and her father is Daniel Charles C.C.O. Motor City Automotive who often use that influence to help Sarah in her advancement but it is her accomplishments that she deserves the credit. With the recent changes in the political climate and the controversies over the study of superhuman biology and certain technology her parents no longer want her working at S.T.A.R. labs but because she is almost 18 there is very little they can do.
Skipping several grades she graduated from Marion High School in Bloomfield Hills Marion at the age of 15. As a result of projects and papers like her study on Cyberware and it's potential emotional impact on society she has won several awards and will be an important part of overseeing Project Red Room once her internship is over.
Powers and AbilitiesEdit
- Medicine
- Leadership


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text:
by MERCEDES JELINEK
These Americans is a series of photographs depicting individuals who attended the Women's March (January 21, 2017) on Washington DC. What began as an internet post after the election of Donald J. Trump, led to over 5 Million people attending marches worldwide; over 500,000 in Washington, D.C. alone. During a time of a polarized America, this day unified demonstrators, gave a voice to those who felt voiceless and fueled what many are calling the 'Resistance'.
*This monograph will be released and shipped in August 2017
*$5/($10 for limited edition) from each book sale will be donated to Planned Parenthood
6 x 9" perfect bound, softcover
72 pages, 40 plates
This monograph is available in two options:
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OPTION II > Book (signed) with 8 x 10" archival pigment print, limited edition of 10 - $100
"I want us to really know us."
Mercedes Jelinek produced these portraits to evoke dialogue about humanity and concepts of identity. Jelinek produced this work so the viewer can take infinite time to study each figure closer, recognize similarities and make connections with the strangers and interactions within each photograph.
Mercedes Jelinek is an artist working in New York. Jelinek is currently a Resident Artist at Penland School of Crafts in North Carolina. She holds a BFA in visual arts from SUNY Purchase College and an MFA from Louisiana State University. Her work has be exhibited globally and has been recently shown at the Ogden Museum of Southern Art, Gatson County Museum, Hinson Art Museum and Blowing Rock Museum.
You will receive one copy of These Americans, signed by Artist Mercedes Jelinek.
Domestic Shipping Only. International Shipping will be available in May 2017. $10 from each purchase will be donated to Planned Parenthood.
This publication ships in May 2017.
UPDATE: Megaphone has SOLD OUT, a new print is available.
You will receive a signed copy of These Americans, along with an 8 x 10" archival pigment print of At the Capitol, 2017. This print edition is limited to ten copies. $10 from each purchase will be donated to Planned Parenthood.
Domestic Shipping Only. International Shipping will be available in May 2017.
This publication ships in May 2017.


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We demonstrate 1.5-W continuous-wave output power from a vertical external-cavity surface-emitting laser (VECSEL) based on InGaAs quantum wells as the gain medium. The VECSEL is pumped by the output of a single-bar diode-laser array at 814 nm and produces an optical-to-optical efficiency of 19%. The high output power is made possible by the use of a sapphire window optically contacted to the intracavity semiconductor surface for heat removal. We demonstrate the good beam quality of the VECSEL output by obtaining 1-W output from a single-mode fiber for 1.5 W launched with simple lenses. Pulsed operation produces a maximum peak power of ∼4.4 W and maximum average power of ∼2 W.
© 2002 Optical Society of AmericaFull Article | PDF Article
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Appl. Opt. 42(33) 6678-6681 (2003)


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Jim Barry Cover Drive Cabernet Sauvignon 2008
Cabernet Sauvignon from Coonawarra, Limestone Coast, Australia
Winemaker's Notes
This wine has a dense purple ruby color with a mulberry hue. The aromas are both opulent and intense with blackcurrant, violets, rosemary and cedar playing a central role. The palate is awash with flavors of boysenberry, cassis, star anise and menthol. The structural tannins are supple and fine grained. Over-all, the wine displays excellent line and length of flavor. An outstanding, youthful, fresh and vibrant Coonawarra Cabernet Sauvignon from a tenanious vintage.
Critical Acclaim
The Wine Advocate - "The fruit for Jim Barry-s 2008 Cover Drive Cabernet Sauvignon comes from 50% Clare Valley and 50% Coonawarra. Deep garnet-purple colored, it gives pronounced aromas of eucalyptus, warm blackberry and crushed blueberries with some cinnamon and cloves plus a whiff of cedar. Medium to high acid and a medium to high level of finely grained tannins support the ample flesh in the mid palate of this full bodied style. The finish is long and minty. Approachable now, it should be best 2011 to 2018. "
Wine Enthusiast - "Representing a good value in top-flight Cabernet, Jim Batty's 2008 The Cover Drive features subtle hints of cinnamon, coffee and vanilla layered over blueberry and plum fruit. It's on the full-bodied side, with plush, velvety tannins balanced by crisp acids. Drink now-2018. Editors' Choice."
Australian Wine Companion - "A truly enjoyable, delicious red berry and cassis-fruited wine, the tannins and oak on the boundary. Drink whenever you wish."
- View All
Jim Barry Winery
From the heart of South Australia, Jim Barry was a legendary and beloved Clare Valley identity. Since 1959, Jim Barry Wines reflect the Barry family's commitment to making table wines with an emphasis on quality and enjoyment. Jim Barry's philosophy of winemaking was very simple: own the vineyards to develop the best fruit flavors possible and retain these flavors during winemaking. The rich, full-bodied Jim Barry wines distinctly embody this simple winemaking philosophy. View all Jim Barry Wines
About CoonawarraView a map of Coonawarra wineries
Located on the Limestone Coast of South Australia, Coonawarra is quite flat, except for the vineyards, which are slightly elevated. That, along with the proximity to the ocean and frequent cloud cover, Coonawarra never gets too hot. There is a small strip of soil, about nine miles long and two miles wide, called Terra Rossa, or red earth. It is the coveted place to plant Shiraz & Cabernet Sauvignon in this small region.
Notable FactsCabernet Sauvignon is the grape of choice here. There is a good amount of Shiraz planted as well, and blends of the two are popular. Cabernet grown in the Terra Rossa shows a sense of place, and the wines are in high demand. The wine is elegant in structure, with prominent acidity and firm tannins, while being full of ripe fruit flavors typical of Australia's style.
About AustraliaLike the United States, which is about the same size, Australia's winemaking regions are huddled into one or two pockets of the country. The state of South Australia, which produces about 60% of the country's wine, also has the most wineries and sub-regions, including McLaren Vale, Clare Valley, Coonawarra and Barossa Valley. New South Wales is home to the Hunter Valley, while the smaller, southern state of Victoria is best known for theYarra Valley. Head way west to the very large state of Western Australia and you'll find the tiny region of Margaret River at the southern tip.
Customer ReviewsSign In to Add Your Review3.53.5 out of 5 stars
19 ratings, 9 with reviews14/23/2012Eduardo Medina - Chula Vista, CA24/3/2017Mr. Marco - East Greenwich, RI35/9/201641/30/2013Very good, nice to have again. Fairly rated and priced.Sparky G - Simsbury, CT310/2/2012I liked it as an every day Cab. I will buy again.49/24/2012
Very nice nose. Smooth and supple. Would buy again!27/13/2012
- Style
- Smooth & Supple
Awful, over ripe and requires a spoon to get through a glass. Typical Aussie crap you get at this price point. Not sure how the critics scored it so high.Retired Wine Guy - Lewes, DE410/1/2011This is a great Cab for the price. I'd really like to give it 4 1/2 stars!412/16/2011
- Style
- Big & Bold
510/28/2011210/15/2011310/13/201139/28/2011Vmiglia - New Baltimore, MI49/17/2011Excellent Wine. Great from start to finish, with an array of flavor for a cab. Ordering more soon! You won't be disappointed.37/28/2011just ok for me.47/6/201156/23/2011
- Pair With
- Cheese > Blue
Great Cabernet from down under.45/31/2011SCKristen - Charleston, SC53/11/2011Was introduced to this wine in a wine class. Recently have not been able to find it locally, so was so excited to order it from wine.com. It was as delicious as I remembered. As my friends and I sat down to a pasta dinner and put our noses in our glasses, we all had the same reaction: "Aaaahhhhs" and "Oh, how I've missed you old friend".
- Style
- Smooth & Supple


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1241 Whiting Street, Wyoming 49509
See more homes for sale in Wyoming, MI
This property has been Sold!
07/06/2016 at $99,000
Great 3 bedroom in great neighborhood. Charming home on corner lots. Lots of character with tray ceilings in living room and formal dining room. Gather the family around the fireplace and enjoy home ownership. Huge kitchen with pantry and large eating area. Tons of storage space, full basement is ready to finish. Second bathroom in basement. Wood floors throughout home. 2 car garage.
Property Information
- Status
- Sold
- Address
- 1241 Whiting Street
- City
- Wyoming
- Zip
- 49509
- County
- Kent
- Township
- City of Wyoming
- Possession
- at close
- Zoning
- Total Finished SqFt
- 976
- Above Grade SqFt
- 976
- Garage Desc.
- 2, Detached
- Water
- Public
- Sewer
- Public
- Year Built
- 1940
- Taxes
- $1,823
Rooms and Land
- MasterBedroom
- 1st Floor
- Bedroom2
- 2nd Floor
- Bedroom3
- 2nd Floor
- Bath
- 1st Floor
- Dining
- 1st Floor
- Family
- 1st Floor
- FormalDining
- 1st Floor
- Kitchen
- 1st Floor
- Other
- 1st Floor
- Basement
- Full Basement
- Heating
- Forced Air, Natural Gas
- Acreage
- 0.1
- Lot Dimensions
- 40 x 115
- Appliances
- Dryer, Oven, Refrigerator, Washer
- Fireplace Desc.
- Family
- Features
- Garage Door Opener, Wood Floor
- Exterior Materials
- Aluminum
- Exterior Features
- Fenced Back, Patio
Listing Video for 1241 Whiting Street, Wyoming MI 49509
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Listing Broker
Listing Courtesy of Keller Williams Realty Rivertown
Office Address 4301 Canal Sw
Listing Agent Stacy Geerts
Listings last updated: . Some properties that appear for sale on this web site may subsequently have been sold and may no longer be available.
Our Michigan real estate agents can answer all of your questions about 1241 Whiting Street, Wyoming MI 49509. Real Estate One, Max Broock Realtors, and J&J Realtors are part of the Real Estate One Family of Companies and dominate the Wyoming, Michigan real estate market. To sell or buy a home in Wyoming, Michigan, contact our real estate agents as we know the Wyoming, Michigan real estate market better than anyone with over 100 years of experience in Wyoming, Michigan real estate for sale.
The data relating to real estate for sale on this web site appears in part from the IDX programs of our Multiple Listing Services. Real Estate listings held by brokerage firms other than Real Estate One includes the name and address of the listing broker where available.
IDX information is provided exclusively for consumers personal, non-commercial use and may not be used for any purpose other than to identify prospective properties consumers may be interested in purchasing.
All information deemed materially reliable but not guaranteed. Interested parties are encouraged to verify all information. Copyright© 2017 MichRIC LLC, All rights reserved.


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prettywoman101's Profile
My name is Irit and I am 49 years old. I leave in the green Galilee region of Israel.
I love creating things, if it's cooking, baking, painting, sculpturing or making jewelry. Sometimes my muse is a material, sometimes a photo I see or a story I read, a leaf on the ground, basically everything surrounds me. My grandmother, who was sawing and crocheting at nights, so she could help raising the family, had a big influence on me for the type of person that she was. She was the one who taught me crocheting.
Like with people, colors that are synchronized make a strong impact on the soul.
Since primary school, the lesson I loved the most was handicraft. When I grow up, I went to the university and graduated Civil Engineering, which is a kind of creation itself, but life brought me back again and again to crafts.
I am married and have two grown up boys. I live in a…
- Female
- Born on July 3
- Joined October 9, 2010
Favorite materials
All kinds of colors, Oil, Acrylic, Pastels, Pencils, Colorful fabrics, Yarn and wires, beads and, Different old materials from which I can make beautiful things
- prettywoman101
- Karmiel, Israel
Woman Beauty


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Ellipse Aura allows acoustic musicians to easily and accurately replicate the sound of their instrument, as mic'd in a professional studio. Players can load up to four Aura Images into this intuitive, discreetly mounted soundhole-mounted preamp. The ergonomic design of the Ellipse Aura puts slider controls at the player's fingertips, helping to maintain a natural playing position.
The Ellipse Aura features Pickup/Image Blend, Volume and switchable Natural I and Natural II Low Frequency toneshaping, a Phase Switch and automatic Anti-Feedback control.
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Bank Of America Q4 2015 Earnings Preview
- Bank Of America customers earn very little from majority of deposit accounts at the bank, partially due to the high switching costs. This is definitely a competitive advantage the bank enjoys.
- The Bank Of America stock is cheap and has plenty of capital to put to use for buybacks or a bigger dividend.
- Long term rates are still falling. Until they rise meaningfully, bank stocks will be rangebound.
If there is one industry that is really forward looking on Wall-Street, it has to be the financial sector. Just look at the performance in the last calendar year, especially that of Bank of America (NYSE:BAC). There was more movement in this stock as a result of sentiment (whether the Fed would move on rates - FOMC meetings) than its earnings results. The stock finished down over 6% despite beating EPS in its second and third quarter. Bank of America Q4 2015 earnings, which will be announced on the 19th of January before the market opens, are estimated to come in at $0.27. Predicted earnings are substantially less than Q3 as business is always slower for this bank in Q4. The spike in volatility (VIX) which is up by 48% since the start of the year hasn't been friendly to the financial sector in 2016. Bank Of America is down over 11% and the financial ETF Financial Select Sector SPDR ETF (NYSEMKT:XLF) is down almost 9%. The S&P 500 as I write is trading at sub 1,900 levels and if it keeps on drifting lower (with elevated volatility), the likelihood of a second interest rate rise in March to me looks slim. However, this stock looks attractive for investors who have confidence in the US economy going forward. If the economy can withstand a tightening cycle by the Fed, Bank Of America's valuation and balance sheet make it one of the stand-out picks in this sector.
Firstly, its balance sheet definitely makes it a leveraged play on rising interest rates and US economic growth. Why? Well, just look at its growing deposit base which came in at $1.162 billion in its latest set of earnings. Now the advantage the bank has here is that 37% of this total (see below) are non interest bearing deposits which means that $429 million carries no interest rate risk to the bank. In effect, whatever interest rates do in the near term, Bank of America will not be more out of pocket (by paying out more interest) from holding this capital which is a huge advantage going forward. Furthermore, on the interest side, the vast majority of this capital is in on-deposit accounts meaning that the interest paid is more or less the same as the Fed fund rate. What does this mean to an investor in this space. Well as long as BAC can continue to pay peanuts on a relative basis on its liabilities (deposits), its net interest income should rise sharply when interest rates rise as its loan book (assets) should become far more profitable and which, incidentally, are growing strongly at present.
Source: Bank Of America Earnings presentation
Bank Of America Stock Trades At Cheap Valuations
The bank's dividend pay-out ratio is only 15% and its debt to equity ratio at a current 1.02 is at its lowest in a decade. If profits continue to flow at the bank, many believe the dividend will get hiked in the near term. This will definitely bring in value investors especially when you take note of the bank's forward price to earnings ratio which has dropped sharply to 8.6 due to recent volatility. Furthermore, the stock is still trading 30% off its present book value which makes the bank one of the cheapest in this sector. Even if the bank prefers to use its excess cash for increasing buybacks (could reach $5 billion in 2016), the stock will still be supported either through internal buying or more value investors coming into the equation.
Nevertheless, its the recent hike in earnings which has re-captivated investor's attention on this stock. The bank has a trailing twelve months EPS of $1.33 which is almost a full $1 in EPS higher than fiscal 2014. If the bank can report an earnings beat next week, it wont be far away from $1.4 in earnings per share for the full year. The market believes momentum will continue as EPS is predicted to get to $1.55 in 2016. However, elevated volatility in markets this year has meant that US bonds have rallied meaningfully. The 20 year treasury bond, for example, iShares 20+ Year Treasury Bond (NYSEMKT:TLT) is up almost 4% which means the 10 year is quickly approaching yields of 2%. Bank Of America is going to find it very difficult to keep earnings growth going if long term interest rates are falling. Bond buying in recent years has been the flight to safety trade. If volatility keeps up and bond buying becomes even more intensified, this could be the perfect excuse for the Fed not to continue with its tightening process.
To sum up, Bank Of America has everything in place to be the US bank that outperforms over the next few years. It has significantly cut expenses, has a strong balance sheet to ensure either strong dividend hikes or buybacks continuing in the future and the stock trades at a low valuation compared to its peers. The one major outlier is the US economy. This bank needs solid loan demand for its deposits, a vibrant housing market and low unemployment to ensure charge off's remain at a minimum going forward. If you are bullish on the US economy going forward, this bank stock will easily return to its present book value per share.


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2008-06-10 at 11:15:47 am #19855http://www.whec.com/article/stories/S471992.shtml?cat=566
XEROX restructuring, but committed to Rochester
officials at XEROX hosted Rochester leaders Monday night to talk about
the company's commitment to the area. During tough economic times, CEO
Anne Mulcahy says XEROX has been able to maintain moderate growth but
admitted restructuring and layoffs are possible.She said, "Rochester
continues to be a place that's attractive to XEROX to have its largest
concentration of people and where we can invest in our toner plant and
Mulcahy and president Ursula Burns say they are
working on a company wide restructuring plan, which could involve
layoffs. They're quick to say any cuts here in Rochester would be
proportionate to cuts made at other areas of operation.Mulcahy said
there are a few main reasons why she will continue to carry a large
part of the work force here. She said, "Part of it's the history. I
would say we have roots. We have 7,600 people, very talented people
that call this home and love it. And quite frankly, we can't blow them
out of here, in terms of their desire to remain in the greater
Rochester area,' said Mulcahy.Last September, XEROX opened a toner
manufacturing plant at the Webster campus. The president says it was
not about staying in Rochester, but what was right for the company.
Burns said because of incentives from the county and state, they built
here. She said, "We do business a lot of different places, employ
people from a lot of different places and so we try to take into
account the hard facts and not the nationalistic facts."
leaders met with area leaders like Assemblyman Joe Morelle Monday
night. He sees XEROX as a success story but believes New York has some
work to do to become more business friendly.Morelle said, "We have to
be mindful. Our cost structure is still too high and we've got to work
on that as a state and region."Anne Mucahy had advice for businesses
struggling here in Rochester. She said, stay focused on your people,
continue to invest in innovation, and listen to your customers.XEROX
has 7,600 employees in the Rochester area. That is more than all the
other US job sites combined.


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The Toffees are still able to finish as high as third with three games left to play and the South African insists they're looking upwards ahead of the Merseyside derby
The Reds trail the Toffees by five points going into the game at Anfield, while Tottenham are only three points ahead in fifth place.
|Steven Pienaar is 11/1 with William Hill to score anytime in an Everton win over Liverpool
"The most important thing is to get into Europe," Pienaar told the club's official website.
"I know at this stage it is difficult but we just want to win the games and see how far we go.
"You don't know what is going to happen on top of us. Maybe one of the teams can lose all their games and we win all ours.
"We just want to win the games that are left."
Everton go into Sunday's game having not won at Anfield since 1999, and a win would guarantee that the Toffees will finish above their neighbours this season and give Evertonians the bragging rights.
He added: "Derbies mean a lot and we know what's at stake.
"For the people of the city and the pride of the club it's one of those games you don't want to lose.
"We will give everything to make sure we come away from Anfield with three points because it has been a long while since we won there."


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Glass fibre tape that is used to support the joining of Topps Tiles construcion boards. This mesh tape comes in a 10m roll and is simple to use and can be cut simply using a pocket knife or scissors, making installation simple.
View our refund and returns policy »
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Christmas Tree Thief Caught By Trail Of Pine Needles
December 19, 2011 by staff
Christmas Tree Thief Caught By Trail Of Pine Needles, A blundering thief who stole Christmas trees from a farm in northern England was caught after leaving a trail of pine needles.
John Dacre was devastated to find his farm in Spenborough ransacked and his pine trees, holly wreaths and festive decorations snatched using his own trolleys, the Spenborough Guardian reported.
But the 65-year-old spotted a long line of green spikes leading away from the farm, and with the help of police officers he followed the trail for a half a mile to a garden where the festive stash was being hidden.
"The funny thing is that they were supposed to be 'low needle drop' trees which don't shed so easily," he said. "But thankfully they dropped enough for us to follow."
A police spokesman said, "The festive foliage led officers to a house where the stolen trees were found and recovered. As an extra present, officers also found a cannabis farm at the address."
A 17 year old was arrested on suspicion of burglary and cultivating cannabis and was bailed pending further enquiries.
Please feel free to send if you have any questions regarding this post , you can contact on
Disclaimer: The views expressed on this site are that of the authors and not necessarily that of U.S.S.POST.


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Contact Our Support Staff
This form will automatically email the Jazz24 support staff. Please fill out all the boxes on the form and provide a detailed message about the issues you are experiencing. We do our best to reply to your message within 24 hours. However, we are not always able to do this depending on the issue you are experiencing or the volume of emails we receive.


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What is a proctored exam?
A proctored exam is administered by an individual who supervises the student while he/she is taking the exam. The proctor's function is to ensure the integrity and security of the exam. You will be notified if your course requires proctoring.
What does the proctor do?
Exams are emailed to proctors on the business day before the exam is to be administered. The proctor prints the exam and gives it to the student. Proctors time the exam, stop the student when the time runs out, and then faxes or scans and emails the exam and any supplementary notes/pages to the OAEE. Once the proctor has the OAEE's written confirmation that the electronic materials were received, the proctor then mails the hard copy to the OAEE. Students will need to be informed if they are to provide postage for the mailing.
Who can be my proctor?
The proctor may be any disinterested third party. A member of the HR/Training staff within your company would qualify to serve in this position. Students have used hotel concierges, librarians, religious leaders (i.e. pastors), and military education officers when an HR/Training staff member is unavailable. The proctor cannot be any person with a vested interest in your success, such as another student in the program, a friend, family member, supervisor, or subordinate. You and the proctor can be in the same department or work on the same project, but one cannot report to the other.
What if I have already have a proctor on file with OAEE?
You must submit a Proctoring Agreement to the OAEE every semester.
Can I come to campus to take my exams with the class?
Please contact your professor for permission to sit with the class on exam dates. You MUST notify the OAEE if permission is obtained.
Are there other local options for proctoring?
- The OAEE proctors exams during regular business hours (8:30 AM – 5:00 PM, Monday-Friday).
- The Distance Education Technology & Services (DETS) office proctors exams after-hours (5:00 PM – 9:00 PM, Monday-Thursday, in 2125 J.M. Patterson).
How do I schedule my exam?
Professors provide the date range for exams to the students. If you are taking an exam outside of the exam schedule, you must provide written permission from the professor to the OAEE, preferably one week in
What is the Honor Pledge?
Information regarding the University of Maryland Honor pledge is here.
If I miss an exam, can I take it later?
If you miss your scheduled exam time, your professor may give you a zero on the exam. When you have a conflict with an exam time/date, please contact your professor as soon as possible (and copy the OAEE) for the permission to take it on another day.
The OAEE reserves the right to deny any proctor or assign proctors to students as necessary. If you have questions about an individual's suitability, contact Brittani Berry, Program Management Specialist, at (301) 405-3017 or e-mail email@example.com.


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King adds to gloom over UK economy
Sir Mervyn King and other Bank of England policy makers warned MPs on Tuesday that the economic situation had deteriorated so severely that they had ripped-up their forecasts of six weeks' ago.
The BoE governor said he was gloomy about the UK or other leading economies staging a robust recovery from the crisis until the eurozone had resolved its problems decisively.
In some of his bleakest remarks to date, coming soon after weak public finance figures, he cautioned the public not to think Britain was even half-way through the crisis, which started almost five years ago.
"When this crisis began in 2007-2008, most people including ourselves did not believe that we would still be right in the thick of it, in the middle of it, quite this late," Sir Mervyn told MPs on the Treasury Committee.
"All the way through, I've said to this committee that I don't think we are yet half-way through – I've always said that and I'm still saying it."
The BoE governor said he had called for more money-printing under the £325bn quantitative easing scheme in June because the situation had deteriorated so far from the monetary policy committee's May forecast.
That had shown a recovery to 2 per cent annualised growth rates by the second half of this year followed by an acceleration in 2013.
"In the last six weeks," he said, "I am very struck by how much has changed since we produced our May Inflation Report."
Describing himself as "pessimistic" about developments in the eurozone, Mr King said, "I have no idea what is going to happen in the euro area". This led him to be unwilling to make what he said would be a "stab in the dark" about when the UK economy could return to normality.
"It is impossible to imagine a situation in which you just do not know what the situation will be in a part of the world that is close to you and is half of your trade. And that makes it impossible to engage in any sensible forecasting."
The chilling effect of the eurozone crisis was wreaking havoc with investment decisions of companies in the eurozone, the UK and in the rest of the world, Sir Mervyn said.
"The confidence that is particularly lacking now is the confidence to invest today rather than wait and see what happens to the euro area and other developments over the next two years.
"It is postponing decisions that companies fully expect to take at some point – that now is not the time to commit. If everyone does that, then you do indeed get a self-fulfilling downturn in the economy."
The only solution Sir Mervyn saw to the crisis was international cooperation in recognising losses stemming from the eurozone and starting to rebuild from there.
"I don't think any one country can get out of this easily on its own. It will require a great deal of international co-operation. And that is why it is very striking that our friends and colleagues in the United States spend so much time worrying about developments in the euro area."
In the meantime, he insisted that the BoE had "not run out of road" with quantitative easing even if he accepted it might not be sufficient to build a self-sustaining recovery.
He also hoped that the new BoE and Treasury funding for lending scheme would soon become operable once it had received state aid clearance from Brussels.
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