text:
Call us to set a party reservation, call ahead seating , or place a To Go Order at
We are located in on the corner of West Poplar St. and North West Ave in Downtown Sidney, Ohio.
Monday — Friday
11:00am – 2:00pm
4:00pm – 9:30pm
Dinner Only
4:00pm – 9:30pm
Welcome to The Bridge!
Accepting Reservations to
Celebrate Mother's Day
at The Bridge, May 8th
Between 11 am – 5:00 pm
On this special day we welcome reservations
for any size group.
Reservations are recommended, please call 937-492-2542
Our Commitment to You
Our goal, create a landmark restaurant that provides a quality dining experience in the heart of Shelby County. Focus on high quality food using premium ingredients, a level of service that meets customer expectations and a fine dining experience in a casual and relaxed manner perfect for all occasions.
About The Bridge.
The Bridge Restaurant was created by Kent Wolters and members of the staff. Kent's interest in history, historic landmarks and developing a close connection with the community made creating the name a pinnacle for our vision. Connecting Kent's interest with the restaurant's name and theme was the challenge.
After many possibilities, it was apparent the Big Four Bridge is a historic landmark that played a significant role in Sidney's economic stability during a time of need in the early 1900's and years following. It was unmistakable the name of the restaurant would be tied to the Big Four Bridge.
Providing a quality dining experience is our commitment. A paramount restaurant in Shelby County will aid further development of Sidney's downtown area and the community. Enjoy.


text:
Dhule riots: Cops on rampage
Posted on Wed , 30 Jan '13
Police in the northern Maharashtra town of Dhule destroyed private property and looted shops earlier this month after violence broke out following a tiff over payment of a hotel bill. About a dozen video clips available with The Indian Express, each running into a few minutes, show the police breaking open a stall, pulling down a pandal meant for hawkers, smashing water drums and pounding two-wheelers parked along the street on January 6. (IE Video)

subset: commoncrawl


text: On Fri, 05 Sep 2008 18:33:07 +0200, Julian Bäume <julian@...> wrote: > Hi! > > On Monday 01 September 2008 13:25:47 00Piano wrote: >> hello there, >> i tried to get ktechlab working on my mac with osx installed (kde, xcode >> tools, qt as well). unfortunatelly, with no success. > After reading your mail a second time, I see a problem here. ktechlab > doesn't > only rely on Qt3 but also on kdelibs3. I don't think that you can run > them on > MacOS. What about this: http://techbase.kde.org/index.php?title=Projects/KDE_on_Mac_OS_X/KDE_3 ? I think it should work.


text:
Birds Schoop Out A Win In The BronxOrioles ace Chris Tillman wins his 14th game of the season
Yankees On A RollMark Teixeira homers in his return to the lineup.Yankees have won the first three games of the series.
Yankees Not Going Away YetThird win in a row puts the Yankees above .500
It Takes 2Bullpen trio holds one run lead
Orioles Have Their Way At FenwayBaltimore takes a one game lead in A.L. East
Sox Bounce BackRed Sox pull back into a first place tie with the Orioles in the A.L. East
Birds Take ChargeLack of run support spoils Price performance
Yankees Nearly Squander 7-run Lead, Beat Orioles 8-6
Birds Rally Past Yankees3 run Yankee lead evaporates
Red Sox BluesOrioles hit a season high 7 home runs
The Mookie Show In BaltimoreBetts homers 3 times and Xander Bogaerts hit streak now at 24 games
Sox Come Up Big In BaltimoreBogaerts extends hitting streak


text:
The 2016 spring season is here!
We now have a new calendar with a range of trips and coaching for everyone and it's available online for you to download.
NCKC 2016 Calendar
Courses will be available for online booking as usual.
The Sunday and Tuesday sessions have started and the weather is starting to get warmer (sometimes) so time to get back on the water, also check out the Events page for up and coming trips and White Water Fridays.
Maybe it's time to get booked onto a 1 or 2 star course – check out what's available here…
If there's something you'd like to try but you can't see it here then get in touch and tell us – it's your club!


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Escondido, California Ssdi Lawyers and Law Firms
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Trust in us to help you win the benefits you are entitiled to. No fees unless you win!
Address: 3737 Broadway Ste. 310 - San Antonio, TX 78209
Phone: (210) 820-3737


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Hilton Garden Inn Sharpsburg hotels are listed below. Search for cheap and discount Hilton Garden Inn hotel rates in Sharpsburg, MD for your personal or business trip. We list the best Hilton Garden Inn Sharpsburg properties so you can review the Sharpsburg Hilton Garden Inn hotel list below to find the perfect place.
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marsh & mclennan cos
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In This Section
Journal Forum
A Meta-Analysis of High Resolution Audio Perceptual Evaluation - June 2016
2 comments
Synthetic Reverberator - January 1960
3 comments
Sound Board: High-Resolution Audio - October 2015
4 comments
AES E-Library
A Unified Fast Algorithm for Cosine Modulated Filter Banks in Current Audio Coding Standards
Current audio coding standards, such as MPEG-I layers 1-3, MPEG-II layers 1-4, MPEG-!V, and AC-3, have adopted for compression various forms of the cosine modulated filter banks (CMFBs). It is demonstrated that all modulated cosine transforms can be divided into two modules - the permutation and discrete cosine transform. The derived discrete cosine transforms are either type II, type III, or type IV. A fast computing algorith is proposed that will compute all three types of transforms uniformly. The new fast algorithm has good features with regard to regularity, complexity, and general applicability.
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text:
Some very juicy programs -- it sounds like a wonderful time!
Travel Tours for Ballet or Dance PerformancesCombined travel tours for attending ballet/dance performances
Posted 16 June 2013 - 10:51 AM
Posted 17 June 2013 - 06:49 PM
Posted 11 October 2014 - 08:29 AM
Posted 30 November 2014 - 09:37 AM
Also, I will soon be planning a trip to Munich and Stuttgart for end of April, and Hamburg ballet days (including guest appearance by Houston Ballet) beginning July. Please get in touch if you are interested in any of these.
Posted 08 January 2015 - 01:39 PM
Posted 08 January 2015 - 07:05 PM
My Munich-Stuttgart ballet trip is now available for booking (deadline Jan 20)
This trip sounds fantastic, Katherine!
Posted 09 June 2015 - 04:56 AM
With the Stuttgart Ballet having announced its season today, I have now completed my list of possible ballet trips for 2016. Please see below. For more info, visit http://toursenlair.blogspot.ca/ or message me.
January 28 - February 1
Florida (West Palm Beach and Sarasota)
Miami City Ballet:
La Source (Balanchine), Barber Violin Concerto (Martins), In the Upper Room (Tharp)
Sarasota Ballet:
Emeralds (Balanchine), new Graziano, TBA
San Francisco
San Francisco Ballet:
7 for Eight (Tomasson), Magrittomania (Possokhov), Pas/Parts (Forsythe)
February 27 - March 4
Washington DC
Mariinsky Ballet:
April 1 - 13
Paris and Amsterdam
Paris Opera Ballet
Seven Sonatas (Ratmansky), Duo Concertant (Balanchine), Other Dances (Robbins), In Creases (Peck)
Romeo and Juliet (Nureyev)
Dutch National Ballet
Apollo, Stravinsky Violin Concerto, Tarantella, Theme and Variations (all Balanchine)
Metaforen, Two, Two, Gold Variations, Adagio Hammerklavier (all van Manen)
April 13 - 20
Berlin and Dresden
Staatsballett Berlin
Multiplicity, Forms of Silence and Emptiness (Duato)
Romeo and Juliet (Cranko)
Dresden Semperoper Ballet
Sleeping Beauty
April 29 - May 10
Prague and Vienna
(day trip to Bratislava)
Czech National Ballet
La Bayadère
Vienna State Ballet
Mayerling (MacMillan)
Marie Antoinette (de Bana)
Hamburg Ballet
Shakespeare Dances (Neumeier)
Slovak National Theatre Ballet
The Sleeping Beauty
New York
New York City Ballet:
Ballo della Regina, Kammermusik No. 2, Vienna Waltzes (all Balanchine)
Dances at a gathering, West Side Story Suite (both Robbins)
American Ballet Theatre: TBA
May 21 - 23 (Victoria Day Weekend)
Boston Ballet
Swan Lake
New Armitage, New Yanowsky, Belong pas de deux (Vesak), Resonance (Martinez)
June 11-20, 10 days
South of France (Toulouse and Bordeaux)
Ballet of the Opéra National de Bordeaux
Messiah (Wainrot)
Ballet of the Théatre du Capitole Toulouse
Paquita grand pas (Petipa), Firebird (Béjart)
July 15 - 25, 11 days
Dancers of the Korean National Ballet, Czech National Ballet, Ballett Zürich, Ballet du Rhin, Augsburg Ballett, Ballett im Revier and Gauthier Dance
Mixed Program
Stuttgart Ballet
Mixed Program (excerpts from Stuttgart Ballet world premieres of the last 20 years)
the second detail (Forsythe), new Goecke, Beethoven's Seventh Symphony (Scholz)
Romeo and Juliet (Cranko)
The Taming of the Shrew (Cranko)
Onegin (Cranko)
John Cranko School Gala
Stuttgart Ballet Gala
Posted 03 June 2016 - 05:55 AM
I have now completed my list of possible ballet trips for 2016-17. Please see below. For more info, visit http://toursenlair.blogspot.ca/2016/06/2016-17-tours-en-lair-ballet-trips.html or message me. You can join a trip from anywhere in the world.
August 10 - 13, 4 days, 2 performances
Saint-Sauveur, Quebec
Guillaume Côté and dancers of the National Ballet of Canada:
The Story of us (Marcelo Gomes, World Premiere, with Heather Ogden)
Venom (Côté)
The Sleeping Beauty prince's solo
Dance me to the end of love (Côté)
- Guillaume Côté – National Ballet of Canada
- Heather Ogden – National Ballet of Canada
- Misty Copeland – American Ballet Theatre
- Blaine Hoven - American Ballet Theatre
- Nehemiah Kish – The Royal Ballet
- Yuhui Choe – The Royal Ballet
- Anne Plamondon - Independent artist
- more dancers TBA
in So Blue
October 14 -16
New York
New York City Ballet
New Lovette, New Peck 1, New Walker, New Lopez Ochoa, Everywhere We Go (Peck)
Dances at a Gathering (Robbins), Firebird (Balanchine)
Glass Pieces (Robbins), Thou Swell (Martins), Stars and Stripes (Balanchine)
January 23 - February 2 (11 days)
Budapest and Vienna
Hungarian National Ballet
Onegin (Cranko)
Vienna State Ballet
La Fille mal Gardée (Ashton)
Cinderella (Malandain)
Carmina Burana (Orlic), Bolero (Lukacs), Afternoon of a Faun (Nebyla)
Ballet of the National Theatre of Brno
Black and White (Radacovsky)
February 3 - 5
San Francisco
San Francisco Ballet
HAFFNER SYMPHONY (Tomasson), JIŘÍ BUBENÍČEK WORLD PREMIERE, IN THE COUNTENANCE OF KINGS (Peck)
SEVEN SONATAS (Ratmansky), YURI POSSOKHOV WORLD PREMIERE, PAS/PARTS 2016 (Forsythe)
February 24 - March 5 (10 days)
Berlin and Dresden (day trip to Leipzig)
Berlin State Ballet
Onegin (Cranko)
Castrati (Duato), Petite Mort (Kylian), Secus (Naharin)
Multiplicity, Forms of Silence and Emptiness (Duato)
Dresden Semperoper Ballet
Swan Lake
Leipzig Ballet
Van Gogh (Schroder)
March 8-12 (5 days)
Sarasota and Fort Lauderdale
Sarasota Ballet
Scènes de ballet (American premiere), The Two Pigeons (both Ashton)
Miami City Ballet
The Fairy's Kiss (Ratmansky), Walpurgisnacht Ballet (Balanchine), Polyphonia (Wheeldon)
April 1 - 12 (12 days)
Zurich and Munich
Zurich Ballet Junior Company
Un Ballo (Kylian), new Favre, new TBA
Zurich Ballet
Swan Lake
(reconstruction of the original Petipa version by Alexei Ratmansky)
Stanislavsky Ballet
Mayerling (MacMillan)
Bavarian State Ballet
Spartacus (Grigorovich)
Symphony in C (Balanchine), In the Night (Robbins), Adam Is (Barton)
Romeo and Juliet (Cranko)
La Fille mal Gardée (Ashton)
Bavarian State Ballet Junior Company
Mixed Program
April 30 - May 10 (11 days)
Amsterdam and Paris
Dutch National Ballet Junior Company
Mixed program
Dutch National Ballet
All Balanchine: Theme and Variations, Tarantella, Stravinsky Violin Concerto, Apollo
Paris Opera Ballet
In G (Robbins), La Valse (Balanchine), Bolero (Cherkaoui)
May 11 -19 (9 days)
An American in Paris (Wheeldon)
Royal Ballet
Mayerling (MacMillan)
The Vertiginous Thrill of Exactitude (Forsythe), Tarantella (Balanchine), Strapless (Wheeldon), new Scarlett
Northern Ballet
Casanova (Tindall)
plus possible other performances by other
companies TBA
Posted 03 June 2016 - 12:42 PM
As always, this looks very juicy!
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text:
Las Vegas Adoption Lawyer, Nevada
Kari J. Hanratty
We understood domestic matters are emotionally driven and as such, out goal is to always listen,... (more)
Emily McFarling Benson
In addition the practice of law, Emily is the author of several children's books and was the operator of an... (more)
Larry C. Johns
Larry C. Johns, a Las Vegas native, received his Juris Doctor from the University of Arizona in 1968 and... (more)
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Dedicated Attorney in Las Vegas, NevadaFamily Law ◊ Real Estate ◊ Timeshare FraudWhen you have... (more)
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Laura Johns is a Las Vegas native and a graduate of the William S. Boyd School of Law at UNLV. She was... (more)
Brian J. Steinberg
Brian is a native of Las Vegas, Nevada. Brian worked in healthcare law and insurance defense before... (more)


text:
The month in review: abstracts from selected press releases and announcements made during the prior month related to web search and search engine marketing.
The links and descriptions below are from official press releases, and may contain marketing hyperbole, claims or euphemisms not necessarily endorsed by Search Engine Watch.
May 5
Google Named Top 5 Business-To-Business Media Property
Google today announced it has advanced to third place in the Media Power 50, an annual review of the most powerful and targeted business-to-business advertising outlets published by BtoB, a leading national marketing publication.
May 6
Espotting Media And Exhibitions.Com Strike Pan-European Marketing Deal
The offline world is being mirrored online. It is traditional for offline exhibitions to partner with media owners to market their events. Now the same premise is coming to the Web, through a pan-European deal between Espotting Media and Exhibitions.com.
May 8
Overture Reports Continued Progress On Growth Initiatives At 2003 Analyst Day
Overture Services, Inc. today reported at its 2003 meeting with financial analysts that the company remains focused on enhancements to its core paid search service, the development of additional search products for syndication to advertisers and distribution partners, and expansion of its international business.
Yahoo And Sprint Make Suite Of Yahoo Services Available To Pcs Vision Subscribers Nationwide
Beginning today, Yahoo and Sprint will make a number of enhanced Yahoo services, with richer graphics and enhanced usability, available to PCS Vision subscribers.
May 13
Dealtime And Epinions Successfully Complete Merger
DealTime, the shopping search engine, today announced that it has successfully completed the acquisition of Epinions, the online consumer reviews and ratings platform. The combined brands rank as the fifth largest online shopping destination.
May 21
Espotting And Kelkoo Expand Deal To Cover 4 European Territories
Espotting Media has expanded its contextual advertising deal with European shopping search engine Kelkoo, to cover a total of 30 channels in 4 European territories – UK, France, Germany and Spain.
Overture Launches in Italy
Overture Services, Inc. today announced the official launch of its Pay-For-Performance search service in Italy. Overture's other non-U.S. business operations include headquarters in Ireland and operations in the United Kingdom, Germany, France, Japan and South Korea.
May 28
Ask Jeeves to Focus Exclusively on Web-wide Search Business with Sale of Enterprise Search Division
Ask Jeeves, Inc. has entered into a definitive agreement under which Kanisa, Inc. will acquire Jeeves Solutions, the enterprise software division of Ask Jeeves.
May 29
Metasearch Engine First To Connect To Over 1000 Online Engines
London based search engine, Turbo10 unveiled technology connecting to over 1000 topic-specific engines on the Internet at The Twelfth International World Wide Web Conference in Budapest last week.
Search Headlines
NOTE: Article links often change. In case of a bad link, use the publication's search facility, which most have, and search for the headline.
Related reading
Four key digital transformation trends you must pay attention to right now
Last week ClickZ Intelligence held its webinar on The What, Why and How of Digital Transformation in association with Marketo.
Google launches imported call conversions
If the internet killed the phone call, the smartphone has revived it.
Will Pokémon GO become a marketing platform for local businesses?
Nintendo's Pokémon Go, a free augmented reality (AR) mobile game, is on fire.
Five most interesting search marketing news stories of the week
Welcome to our weekly round-up of all the latest news and research from around the world of search marketing and beyond.


text:
Nerve wants to be a cautionary tale about the perils of desiring fame through social media, but it isn't willing to go to the darker…
Nerve wants to be a cautionary tale about the perils of desiring fame through social media, but it isn't willing to go to the darker…
The Star Wars-ification of Star Trek continues; better than the others, but still not good enough.
"The Ballad of Narayama" is a Japanese film of great beauty and elegant artifice, telling a story of startling cruelty. What a space it opens…
Patrice Leconte's "Monsieur Hire" is a tragedy about loneliness and erotomania, told about two solitary people who have nothing else in common. It involves a…
Ken Loach's "I, Daniel Blake" wins the Palme d'Or at the 69th Cannes Film Festival.
Barbara Scharres on the cats of Cannes 2016.
"It's a madhouse!"
Wednesday, July 18, is the 20th anniversary of our marriage. How can I begin to tell you about Chaz? She fills my horizon, she is…
Writers at RogerEbert.com pick their favorite movies featuring star-crossed lovers.
Writers at RogerEbert.com pick their favorite movies featuring aliens and UFOs.
Netflix's new docu-series tackles a fascinating story of a community college that serves as the final stop for dozens of potential superstars.
A Comic-Con 2016 report from the "Creating Universes" panel, featuring Neil deGrasse Tyson, Len Wein, Bill Prady and Peter S. Beagle.
* This filmography is not intended to be a comprehensive list of this artist's work. Instead it reflects the films this person has been involved with that have been reviewed on this site.
A recap of the 2016 TCM Classic Film Festival.
A report on the 2016 Film Independent Spirit Awards.
A list of the 2016 Academy Award nominees.
An article about the 2015 Alliance of Women Film Journalists' EDA Award Winners.
A preview of the 73rd Golden Globes ceremony airing Sunday night, and some predictions.
Highlights of our 2015 interviews, including Brie Larson, Bryan Cranston, Jason Segel, Lexi Alexander, Sarah Silverman, Spike Lee, Tom McCarthy, Ramin Bahrani, Paul Feig, Charlie Kaufman and much more.
Contributors to RogerEbert.com each list their favorite films of 2015.
The ten best films of 2015.
A news story on the 2015 CFCA nominees.
An article on the 2016 Golden Globe nominees.
An interview with co-writer/director Tom McCarthy about "Spotlight."
An overview of the films that will be theatrically released in the 2015 fall season.
Marie writes: They call it "The Shard" and it's currently rising over London akin to Superman's Fortress of Solitude and dwarfing everything around it, especially St. Paul's in front. I assume those are pigeons flying over-head and not buzzards. Ie: not impressed, but that's me and why I'm glad I saw London before they started to totally ruin it.Known as the "London Bridge Tower" before they changed the name, when completed in 2012, it will be the tallest building in Europe and 45th highest in the world. It's already the second highest free-standing structure in the UK after the Emley Moor transmitting station. The Shard will stand 1,017 ft high and have 72 floors, plus another 15 radiator floors in the roof. It's been designed with an irregular triangular shape from base to top and will be covered entirely in glass. The tower was designed by Renzo Piano, the Italian architect best know for creating Paris's Pompidou Centre of modern art with Richard Rogers, and more recently the New York Times Tower. You can read an article about it at the Guardian. Here's the official website for The Shard. Photograph: Dan Kitwood.
Sun-Times Gallery of Top Oscar Categories
Sun-Times Gallery of Top Oscar Categories
Based on his show-stopping speech at Saturday night's Independent Spirit Awards, if Mickey Rourke wins an Oscar on Sunday night the Oscarcast is going to be a lollapalooza. As his comeback film "The Wrestler" won for best film, male actor and cinematography, Rourke brought the show to a halt and the audience to its feet with an acceptance speech that was classic Mickey. The Indie Spirits are telecast live and unbleeped, which added considerably to the speech's charm.
A man who has received a large sum of money hires people to re-enact scenes from his own life, staged on the actual locations and on sets he has constructed for the purpose.
That's a selective synopsis of the premise of "Remainder," a 2005 novel by Tom McCarthy. As I was sitting through "Synecdoche, New York," I couldn't help feeling that I'd somehow seen this done before (yeah, I know -- the movie is in part about that feeling)... and then I remembered "Remainder." The first-person narrator, who has suffered brain damage in an accident, becomes obsessed with meticulously reconstructing the events surrounding it. Having turned his apartment building, and the blocks around it, into a living set -- available round the clock for command performances, he stages a run-through of one sequence in a warehouse at Heathrow:
I'd had a raised viewing platform built, a little like an opera box, because I'd enjoyed watching the action in my building from above and wanted a similar option here. I'd established that I might roam around the re-enactment area itself, and that the re-enactors shouldn't be put off by this. I chose to begin watching the re-enactment from the platform, though.
Later, he describes his living role as actor, director and audience, revising and perfecting the re-enactment, which becomes a more-or-less permanent project:


text:
In the 1980s, General Motors learned a difficult lesson about technology when it copied Japanese auto manufacturers in their use of robotics — but didn't copy anything else. Thinking that large capital investments in technology were the key element to the success of Japanese automotive supply chains, GM spent billions purchasing hundreds of industrial robots. Management claimed the increased automation would significantly boost productivity. Instead, the robots broke down and started painting themselves, breaking windshields and even welding doors shut at a Cadillac plant. Yet beyond the production delays and the expensive repairs required to straighten out the wayward robots, GM missed an essential point. The Japanese automakers saw robots as just one element of a larger supply chain design. GM's competitors were successful not because of a technological silver bullet but because they had designed an operating plan that translated customer-focused product design into a simplified, flexible, self-improving supply chain. The lesson? There are no quick fixes or cure-alls. You need a comprehensive approach. Download this paper to learn how!
E-Signatures and Digital Signatures: 10 Things Procurement Needs to Know
From better contract management and accelerating the flow of the cash cycle to simply boosting productivity, electronic and digital signatures have proven to be a useful tool for procurement organizations. And, as the supply chain — and the rest of the world — continue to move toward a paperless workflow and implement smarter technologies, digital signatures and e-signatures are a natural path or progression for procurement. Bottom line: digital signatures and electronic signatures save time and money. Download this paper to learn more!
Total Talent Management and the Contingent Workforce Paradox
Spend Matters - Free Research Download
Organisations realize they must consider the contingent workforce similarly to how they manage internal staff — in the search for talent, in managing performance and even in retention or exit. At the same time, the contingent workforce is different, and thus requires different processes and management, from the permanent group. The current regulatory environment, while not the only driver, means that governments will increasingly require careful management of the contingent workforce if organisations are to stay within the law and avoid reputational and financial risk.
VMS and Services Procurement Technology Selection in 2015 — New Tools, Shifting Strategies, False Prophets and More
Spend Matters - Webinar Replay
If you are thinking about implementing a VMS or other services procurement technology solution (e.g., freelancer management system), you can't afford to miss this informative session.
Applying Supply Chain Rigor to Contingent Workforce Management
In today's global economy, firms like Amazon, Apple, GE, P&G, and Walmart all compete on how well they manage their extended supply chains and associated spending on critical goods and materials. But, in an increasingly services-based economy where nearly everything is becoming a "service" (including all the non-production elements associated with creating a product), applying the same rigor to managing external services spending should also be important. What are the main differences with the contingent market? Why does it matter? What are your peers saying about the space? Download this survey-based report to find out!
Relationship Counseling for Staffing and Services Procurement – Successful Therapy For Lasting Partnerships
There are many reasons for the uniqueness of the services procurement ecosystem within procurement; this "uniqueness" can vary dramatically by industry and region as well. But a universal theme we've observed for years is a significant disconnect between buyers and suppliers — especially between the corporate procurement function and the vendor community (even if the relationship between the non-procurement line of business manager and the vendor is just fine). So how do you bridge the disconnect? Download this paper to find out!
Procurement Consultants: Sourcing, Implementation, and How to Get the "A" Team
Spend Matters - Free Research Download
Over the years, we've come to realize what a significant disconnect there is between those on the inside of the consulting world and those who buy services, especially in the operations and procurement areas. If you're looking to hire Mr. T in this regard, we unfortunately can't help you. But we can tell you how to get the most out of a consulting team.
VMS and Services Procurement M&A: Customer Tips and Tactics
Spend Matters - Free Research Download
Spend Matters believes that there's a 90% chance that one of the top vendor management system (VMS) players in the market (Fieldglass, IQNavigator, or Beeline) will be acquired within the next year. It's important for customers and competitors to know how to properly prepare for acquisition activity in the services procurement sector. This paper offers actionable tips and strategies to help procurement professionals get smart, with generic software and VMS-specific tips.
How to Attack Marketing Spend
Spend Matters PRO Only - Premium Research Download
One of our most popular topics on PRO, Thomas sets out to define how procurement should approach the marketing category (from dealing with agencies all the way to just dressing better!) For a category that straddles the boundary between direct and indirect spend, this is an indispensable guide for both procurement and marketing departments.
Procurement's View of Outsourcing
Spend Matters PRO Only - Premium Research Download
Spend Matters UK/Europe's Executive Editor Peter Smith tackles "cases where an activity has been performed inside the organisation, at least in part, and is then transferred to delivery via an external contractual relationship," all from procurement's perspective of course. He suggest critical factors to pay attention to throughout the entire outsourcing process and solid recommendations for success.
School Catering: Value for schools
Spend Matters Case Study
This paper looks at how optimisation tools are changing the way in which organisations can achieve increased value and performance from large-scale and complicated sourcing challenges.


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Extended Stay America San Francisco - Belmont, Belmont
- Free continental breakfast, free WiFi, and free parking
Main amenities
- 108 smoke-free guestrooms
- Coffee/tea in a common area
- Air conditioning
- Front desk safe
- Self-serve laundry
- Limited hour front desk
For families
- Kitchen
- Cookware, dishware, and utensils
- Stovetop
- Microwave
- Refrigerator
- Private bathroom
What's around
- San Francisco International Airport
- Peninsula Museum of Art (1.1 mi / 1.8 km)
- Hiller Aviation Museum (1.5 mi / 2.4 km)
- Museum of San Carlos History (2 mi / 3.1 km)
- Hillsdale Shopping Center (2.7 mi / 4.4 km)
- San Mateo County Event Center (3.1 mi / 5 km)
Extended Stay America San Francisco - Belmont, Belmont
Hotel Info: 020 3027 9803from £118
- Studio One Queen Accessible Smoking
- Studio One King Accessible Non Smoking
- Signature-Studio King (NSmk&Accessible)
- Studio Two Doubles Accessible Non Smokin
- Studio One Queen Accessible Non Smoking
- Studio King (Smoking)
- Studio King (NonSmoking)
- Studio One Queen (NonSmoking)
- Signature-Studio 2 Doubles (NSmk&Acc)
- Studio Queen (Smoking)
- Studio Two Doubles (Smoking)
- Studio Two Doubles (NonSmoking)
Also popular with travellers from the UK
At a glance
Key facts
Hotel size
- This hotel has 108 rooms
- This hotel is arranged over 5 floors
- Check-in time 3 PM-midnight
- Check-out time is 11 AM
Required at check in
- Credit card deposit required
- Government-issued photo ID required
Travelling with others
- Children (17 years old and younger) stay free when occupying the parent or guardian's room, using existing bedding
- No cribs (infant beds)
- Pets allowed *
- Restrictions apply *
- 2 per room
- Free WiFi in public areas
- Free WiFi in rooms
- Free self parking
Other information
- Smoke-free property
Nearby Two Star Belmont hotels
In the hotel
- Free continental breakfast daily
- Coffee/tea in a common area
- Use of nearby fitness center (discount)
- Sailing nearby
- Windsurfing nearby
- Front desk (limited hours)
- Laundry facilities
- Year Built 2003
- Elevator/lift
- Safe-deposit box at front desk
- Designated smoking areas
In the room
- Air conditioning
- Coffee/tea maker
- Iron/ironing board
- Blackout drapes/curtains
- Private bathroom
- Shower/tub combination
- Free toiletries
- 32-inch flat-screen TV
- Premium TV channels
- Desk
- Free WiFi
- Free local calls
- Refrigerator
- Microwave
- Kitchen
- Stovetop
- Cookware, dishware, and utensils
- Weekly housekeeping provided
- Access via exterior corridors
Top Belmont hotels
Recently reviewed hotels in Belmont
- Adante Hotel, a C-Two HotelFrom
"We stayed here for 4 nights in June. It was a great base to stay and everything is within walking distance. "Jun 17, 2016
- Pier 2620 Hotel Fisherman's WharfFrom
"Great hotel fab stay for 12 nights , found the hotel in just the right spot to gain access to everything you could wish for , will be back again ..."Jun 17, 2016
- Stanford Court San FranciscoFrom
"Very comfortable hotel, bed was really nice. Make sure you can walk well if you stay here as at the top of a very steep hill..."Jun 14, 2016
- Oceano Hotel and SpaFrom
"Sent last night of a business trio in the valley here to get aware from the city areas. Much more pleasant and 30 minutes from SFO."Jun 13, 2016
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Extended Stay America San Francisco - Belmont, Belmont's small print
Also known as
- Extended Stay America San Francisco Belmont Hotel
- Extended Stay America San Francisco Belmont
- Extended Stay America San Francisco Belmont Hotel
A valid credit card is required at check-in for incidentals/damages otherwise a US$100 cash deposit per stay, per room will be collected. The deposit is refunded at check-out after satisfactory inspection. If refund is over US$100 a check will be mailed within 14 days of departure.
Pet-friendly rooms can be requested by contacting the property at the number on the booking confirmation.
Optional extras
Weekly housekeeping is provided; fees apply for additional cleaning
Pets are allowed for an extra charge of USD 25 per pet, per day (maximum USD 150 per stay)
We have included all charges provided to us by this hotel. However, charges can vary, for example, based on length of stay or the room you book.
Recent reviews
Good 3.3 /5 from 108 reviews
Great walking area for my 13 lb dog. Cost was $25/night for her--what are you going to do? The majority of properties in the area don't even accept dogs. Staffing is a little short with just one desk person. Often a sign on the desk reads they are with another customer and are not available for customer service. Free breakfast is very minimal, to the point that there's no orange juice or milk for cereal (only hot instant oatmeal). If these two liquids are not available, one cannot call it breakfast.
Hard to get pet in and out on second floor and the distance into the heart of San Francisco was longer than expected.
Better than good. Price fair for area. Neighborhood so, so. Customer Service 100%. CONS - bad pillows, no coffee pot for early and late folks. no cups for coffee or tea. no silverware or plasticware to use in studio kitchen. limited, limited, limited breakfast.
Nice hotel with grab and go breakfast. Convenient to the 101. Allows pets for a fee.
We were able to get a room even tho we called just hours earlier. We came in late and discovered we forgot a hairdryer. Of course they had one! Definitely added it to our short list of good spots to stay
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Vision Statement: All San Jose Unified School District students are inspired and prepared to succeed in a global society.
Mission Statement: San Jose Unified School District's mission is to eliminate the opportunity gap and provide every student with the finest 21st century education.
You are encouraged to attend an AP Bootcamp to learn vital study skills necessary to be successful in your upcoming AP class. This camp is especially useful for students who are taking their first or second AP course.
AP Camp—-August 9-11, 2016
9am to 11 am
WGHS Mickey Long Library
We are getting ready for new murals this year at WGHS thanks to a generous grant being put together by District 6 representative Pierluigi Oliverio. Check out these concept sketches from the muralist.
Now is the time to book your senior portrait appointment. Please visit http://www.stilllightstudios.com/Schools/Willow-Glen-High-School/Home to book your appointment ASAP There are two summer dates at WGHS for photos and there is no sitting fee. You don't want to miss this opportunity to book early and ensure your photo is in the yearbook. Reserve your appointment now!
Summer Hours
Open from 9am to 2pm
Monday through Friday
6/6/16 through 8/12/16
*except during maintenance
The offices and phone lines will be closed for maintenance from 6/27/16 through 7/8/16.
Freshmen - Link Crew Orientation (schedules are given to 9th graders as part of the orientation) - Thursday, 8/4, 8am to 12pm, in the gym. Freshmen are expected to be present for the entire orientation
Regular Schedule pick up for 10th through 12th graders - Wednesday, August 17th, beginning at 6:45am, on the first day of school
Early Schedule pick up opportunity - Wednesday, 8/3
-10th graders, 8:00-9:30am, Wednesday, 8/3
-11th graders, 9:30-11:000am, Wednesday, 8/3
-12th graders, 11:00-12:00pm, Wednesday, 8/3
-Ramp Up to Geometry - 6/7-7/6, M-F, 7:30-2:00
-Ramp Up to Algebra - 6/7-6/28, M-F, 8:00-12:00
-Summer School Session 1 - 6/6-6/24, M-F, 8:00-12:30
-Summer School Session 2 - 6/27-7/18 (closed 7/4), M-F, 8:00-12:30
Congratulations to our NCAA Student-Athlete Commitments. From left to right, Elan Hawkins (Academy of Arts San Francisco - Golf), Ronnie McDowell (Bethune Cookman University - Golf, and Nate Maggi (Fresno State University - Baseball). Not pictured Caleb Vander Esch (South Dakota University - Football)
Congratulations to Willow Glen's Anthony Navarro and Clayton Wilson for being selected to the Charlie Wedemeyer All Star South team.
The 42nd Annual Charlie Wedemeyer All-Star Football Game, presented by the Rotary Club of Almaden Valley, will be held at Levi's® Stadium on Saturday, July 16, 2016. The event is scheduled for 6:00pm kickoff and will feature the very best graduating seniors from 46 Santa Clara County High School football programs. The next generation of collegiate and pro hopefuls will lace up for a special evening at Levi's® Stadium to raise money for charities supported by the rotary club. Over the years, the Rotary Club of Almaden Valley, a nonprofit organization, has raised hundreds of thousands of dollars for charities both local and abroad. The Charlie Wedemeyer game accounts for a great deal of the funds generated by the organization each year.
Interested in what's happening with our WGHS graduates? Click here to see, and send us your great stories.
©2016 Willow Glen High School|2001 Cottle Ave. San Jose,|San Jose, CA 95125| Tel: 408.535.6330 FAX 408.535.2353 Attendance FAX 408.267.5757
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Non-Discrimination Statement
San José Unified School District does not discriminate on the basis of age, sex, sexual orientation, gender, gender identity, gender expression, ethnic group identification, race, ancestry, national origin, religion, marital or parental status, color, mental or physical disability, or on the basis of a person's association with a person or group with one or more of these actual or perceived characteristics in its educational programs and activities or employment practices as required by Americans with Disabilities Act (ADA), Title VI of the Civil Rights Act of 1964, Title IX of the Education Amendments of 1972, the Age Discrimination Act of 1975, Section 504 or the Rehabilitation Act of 1973, and the Vocational Education Act of 1976. The lack of English language skills will not be a barrier to admission and participation in the District's program s. Students, parents, employees, or others who wish further information about these regulations, or who wish to file a complaint, should contact the following persons: Director of Student Services, Dane Caldwell-Holden at 408-535-6080 ext. 13212; Assistant Superintendent of Human Resources, Dominic Bejarano at 408-535-6139 ext. 15015; Manager of Academic Supports, Michelle Herlth at 408-535-6008 ext. 14314; SJUSD 504 Coordinator, Yvonne King at 408 535-6195 ext. 13229.
Declaración contra la discriminación
El Distrito Escolar Unificado de San José no discrimina por motivos de edad, sexo, orientación sexual, género, raza, identificación de grupo étnico, linaje, religión, color de piel o discapacidad física o mental, en sus programas y actividades educativos ni en sus prácticas de empleo, de acuerdo con los requisitos de la Ley de Estadounidenses con Discapacidades (ADA), el Título VI de la Ley de Derechos Civiles de 1964, el Título IX de las Enmiendas a la Educación de 1972, la Ley de Discriminación por Edad de 1975, Artículo 504 o la Ley de Rehabilitación de 1973 y la Ley de Educación Vocacional de 1976. La falta de destrezas en el idioma inglés no será una barrera para ser admitido y participar en los programas del Distrito. Los estudiantes, padres de familia, empleados u otras personas que deseen más información acerca de estos reglamentos o que deseen presentar una queja, deben ponerse en contacto con las siguientes personas: Director de Servicios Estudiantiles, Dane Caldwell-Holden, 408-535-6080 ext. 13212, Superintendente Auxiliar de Recursos Humanos, Dominic Bejarano, 408-535-6139 ext. 15015, Administradora de Apoyo Académico, Michelle Herlth 408-535-6608 ext. 14314, Coordinadora del Artículo 504 del Distrito, Yvonne King 408 535-6195 ext. 13229.


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Conceived and designed the experiments: XZ SH ISAM JY. Performed the experiments: YF YY LP. Analyzed the data: YF HW TZ. Contributed reagents/materials/analysis tools: MY XY. Wrote the paper: YF HW TZ.
Based on next-generation sequencing data, we assembled the mitochondrial (mt) genome of date palm (Phoenix dactylifera L.) into a circular molecule of 715,001 bp in length. The mt genome of P. dactylifera encodes 38 proteins, 30 tRNAs, and 3 ribosomal RNAs, which constitute a gene content of 6.5% (46,770 bp) over the full length. The rest, 93.5% of the genome sequence, is comprised of cp (chloroplast)-derived (10.3% with respect to the whole genome length) and non-coding sequences. In the non-coding regions, there are 0.33% tandem and 2.3% long repeats. Our transcriptomic data from eight tissues (root, seed, bud, fruit, green leaf, yellow leaf, female flower, and male flower) showed higher gene expression levels in male flower, root, bud, and female flower, as compared to four other tissues. We identified 120 potential SNPs among three date palm cultivars (Khalas, Fahal, and Sukry), and successfully found seven SNPs in the coding sequences. A phylogenetic analysis, based on 22 conserved genes of 15 representative plant mitochondria, showed that P. dactylifera positions at the root of all sequenced monocot mt genomes. In addition, consistent with previous discoveries, there are three co-transcribed gene clusters–18S-5S rRNA, rps3-rpl16 and nad3-rps12–in P. dactylifera, which are highly conserved among all known mitochondrial genomes of angiosperms.
The widely-accepted hypothesis about the origin of the mitochondrion assumes that it descended from an endosymbiontic event involving an α-proteobacterium-like organism and the common ancestor of eukaryotes [1]. Evolving from algae to land plants, including bryophytes and angiosperms, plant mitochondrial (mt) genomes have increased their sizes, especially in the non-coding region. Among land plants, bryophytes, i.e., liverworts, mosses, and hornworts, represent the basal forms. They have similar gene order, genome size, and a fraction of non-coding sequences [2], [3]. As evolution continues, land plants gained new mechanisms to facilitate frequent gene exchange between mitochondrial and chloroplast genomes as well as between mitochondrial and nuclear genomes [4], [5]. For instance, mitochondrial genomes of angiosperms have long been known for their slow evolutionary rate [6], existence of subgenomic circles in addition to a master genomic circle [7], extraordinarily large and highly variable genome sizes [8], trans-splicing of group II introns [9], high density of RNA editing [10], [11], divergent non-coding sequences [12], and frequent gene transfer [13]. The inter-genomic gene transfer, together with the continuing increase of non-coding DNA sequences, leads to a broad size range in angiosperm mt genomes, which as of today is from ~200 to 2400 kb based on the known mt sequences and experimental estimations [8], [14]; up to date (July, 2011), there have been more than 40 plant mt genomes sequenced, including 22 angiosperm mt genomes (http://www.ncbi.nlm.nih.gov/genomes/GenomesGroup.cgi?taxid=33090&opt=organelle).
Phoenix dactylifera L., also known as date palm, is economically the most important plant in the Middle East and North Africa [15], and it is estimated to have more than 450 cultivars or varieties in the Kingdom of Saudi Arabia and nearly 2,000 varieties around the world [16]. Therefore, sequencing its mitochondrial genome, together with its nuclear [17] and chloroplast genomes [18], is of essence in improving its agricultural, horticultural, and nutritional values. In this study, combining data from two next-generation sequence platforms, pyrosequencing (Roche GS FLX) and ligation-based sequencing (Life Technologies SOLiD), we assembled P. dactylifera mt-genome (cultivar Khalas, Al-Hasa Oasis, Saudi Arabia) –the first from the Arecaceae family. In addition, analysis of the mt genome sequence and transcriptomic data are of importance in revealing mechanisms underlying mitochondrial genome evolution and the unique evolutionary status of P. dactylifera among angiosperms. Furthermore, based on the data from three commonly-grown cultivars, we also investigated RNA editing sites and SNPs within the species.
We assembled the P. dactylifera master mt chromosome into a 715,001 bp circular molecule (Figure 1; the assembling details are described in Materials and Methods) with an average GC content of 45.1%; it is now the fourth largest mt genome sequenced after those of Cucumis sativus (1,555,935 bp) [19], Cucurbita pepo (982,833 bp), and Vitis vinifera (773,279 bp). Its protein coding sequence is composed of only 6.5% of the genome (46,770 bp) and this gene content is similar to other published angiosperm mt-genomes (Table S1 and Table S2). The rest, also the majority (93.5%) of the genome, is composed of non-coding (the cp-derived regions are also considered as non-coding in this regard), which harbors 0.33% tandem and 2.3% long repeats (the repeat lengths are greater than 50 bp). RNA genes and intron sequences are 1.1% and 4.3% of the mt genome, respectively. This mt genome also contains the second highest proportion (10.3%) of cp-derived sequences among the sequenced mt genomes to date, of which several intact genes, such as petA, petG, petL, psaJ, psbT, rpl20, rpl33, and rps8 are identified. Since the age of the cp-derived sequences or time when the sequences inserted into mt genomes varies greatly [20], we are unable to prove whether these genes are actually transcribed or active since we extracted the total RNA (contains both nuclear and organellar transcripts) from each tissue for constructing transcriptomic libraries among which the expression data of the cp-derived sequences and authentic cp sequences are impossible to separate (see Materials and Methods for more details). Most of P. dactylifera mt genome is rather diverged from other angiosperms. For example, only ~21% of P. dactylifera mt genome sequence is shared (over 70% identity) by Vitis, Oryza and Bambusa, and even less by Zea (~15%) and Arabidopsis (~11%). In addition, consistent with results from previous studies, we observed that three co-transcribed gene clusters, 18S-5S rRNA, rps3-rpl16, and nad3-rps12, are conserved in other angiosperm mt genomes [21]. We summarized general mt genome features including size variations, AT content, and intron types of 15 non-redundant sequenced plant mt genomes (including 12 higher plants and three lower plants) in Table 1. Our phylogenetic analysis based on 22 concatenated conserved genes among 15 selected mt genomes (Figure 2) revealed that P. dactylifera appears to be the more basal among monocots.
The P. dactylifera mt genome contains at least 38 protein-coding genes and five complete ORFs, most of these genes encode proteins of the electron transport chain, such as nine subunits of nicotinamide adenine dinucleotide dehydrogenase (complex I), apocytochrome b (complex III), three subunits of cytochrome c oxidase (complex IV), five subunits of ATP synthase F1 (complex V) and four cytoplasmic membrane proteins required for cytochrome c maturation (Table 2).
We compared these protein coding genes to 11 other sequenced angiosperm mt genomes (Table S2). First, P. dactylifera mt genome does not have the genes encoding respiratory chain complex II, such as sdh3 and sdh4, which are only found in two dicots, Nicotiana tabacum and V. vinifera. Second, our assembly is similar to V. vinifera, and both contain one copy of RNA polymerase gene harboring a conserved domain characteristic of pfam00940 superfamily of polymerases [22]. Third, rps14 present in Brassica napus and V. vinifera is also found here, whereas rps11, another ribosomal protein gene, is exclusively detected in our assembly. Both genes have full open reading frame (ORF) and are likely functional in date palm, though in many other known angiosperm mt genomes they are either pesudogenes or transferred into nuclear genomes [23], [24]. Fourth, the recently identified rpl10 gene, being identified as orf-bryo1 in vascular plants and charophycean green algae [25] and orf168-related sequences in bryophytes and angiosperms [26], seems to be interrupted in our assembly and possibly because of a frame shift event. Fifth, we found several pseudogenes in our assembly, which appear intact in other mt genomes, such as orf99-b (as orf100-ψ in our gene list) in Zea mays and cp-derived gene psbT in V. vinifera. In addition, some of the universally expressed ribosomal genes, including three rRNA genes (5S, 18S, and 26S ribosomal RNA genes), are also unambiguously identified [27]; 5S and 18S rRNA genes are also closely related and distant from 26S rRNA gene in date palm mt genome.
A genome-wide screening yielded 30 full-length tRNA sequences (Table S3) in our assembly; among them, 12 seem to be cp-derived, which exhibit higher sequence identity (>98%) to their chloroplast counterparts than their mitochondrial counterparts [18], and three predicted tRNAs seem to have introns. Among these 30 tRNA genes five amino acids (A, L, R, T, and V) are not encoded, although tRNAs for 20 amino acids are necessary for protein synthesis in mitochondria. In addition, having compared the date palm tRNA gene content to those of seven other plants mt and cp genomes (Figure 3), we conclude that there are 10 tRNA genes, among which nine encoding tRNAs for the five amino acids, are actually lost after the divergence of liverworts from seed plants. These results suggest that the missing tRNAs are supplied by either the chloroplast or nuclear genomes. In addition, we found that four mt tRNA genes of higher plants are gradually lost and replaced by cp-derived tRNA [28]. The reason why mt tRNA genes are replaced by both cp-derived and nuclear counterparts remains an open question. There is also another possibility–all mt tRNA genes may eventually be replaced and what we observed here is only an intermediate and dynamic process.
Chloroplast and mitochondrial genomes are known to share sequences due to frequent gene transfer events [5], [29], [30]. Frequent DNA transfer from cpDNA to mtDNA occur as far back as the common ancestor of the extant gymnosperms and angiosperms, about 300 MYA (million-years-ago) [20]. Our assembly contains more than 100 fragments of chloroplast origin (over 80% identity), ranging from 50 to 6,521 bp in length (Table S4). The total fraction of chloroplast DNA sequences present in P. dactylifera mt genome is 73,691 bp, corresponding to 10.3% of the whole mt genome, and 46.5% of P. dactylifera cp-genome. The proportion of cp-derived sequences in our mt genome assembly is comparable to the two large sequenced plant mt genomes, V. vinifera (8.8%) and Cucurbita pepo (11.6%) [31], but larger than those of the other known plant mt genomes (Table 1). These results suggest that chloroplast DNA sequence insertion is an important mechanism for plant mitochondrial genome size expansion and sequence diversity.
Most of chloroplast sequence insertions in P. dactylifera mt genome are unique, as evident from the observation that only nine out of 44 insertions (over 200 bp) have full-length homologous sequences shared by other known mt genomes (>90% length coverage, >70% identity) (Table 3). Among the nine cp-derived homologous sequences, six and five of them are also found in Vitis and Bambusa, respectively, whereas none is found in A. thaliana. These nine insertions tend to have higher GC content, resembling that of mt genomes as compared to the unique and possibly new insertions (Figure S1), which suggests that these cp-derived sequences did, in some extent, gradually increase their GC content to become similar to their host mtDNAs.
We identified 23 group II introns in 10 protein-coding genes, including four trans-spliced introns of nad1 and nad5, and 20 cis-spliced introns in ccmFc, cox2, nad1, nad2, nad4, nad5, nad7, rpl2, and rps3. No group I intron was discovered in our assembly. In general, the functional mitochondrial rRNA and tRNA genes of the sequenced angiosperm mt genomes do not possess introns, but we found three intron-containing tRNA genes in our assembly: trnK-TTT, trnN-ATT and trnSup-CTA, and we have yet to validate if they are functional or not.
Mitochondrial RNA editing is essential for functional protein synthesis since nearly all plant mt mRNAs are edited [32], [33], [34] and it modifies amino acids and generates new start or stop codons [35], [36], [37], [38], [39], and it has been documented in most plants except algae and mosses. It suggests that this cellular process is ancient arisen in early land plants after they split from Bryophyta [10]. We predicted nearly 600 putative RNA editing sites (Table S5) using PREP-Mt [40]–an effective tool identifying C-U editing sites. We found that the nad4 gene contains the most edited sites (59). In addition, our comparative analysis revealed that 305 (51.5%) and 278 (47.0%) C-U editing sites in date palm are shared by O. sativa and A. thaliana, respectively (Figure 4). Experimental examination confirmed 40 of 41 predicted C-U editing sites in five randomly chosen genes (atp1, atp4, atp9, rpl116 and rps19) using cDNA sequences (Table S6) and additional nine sites not detected by PREP-Mt were identified. We also compared their tissue disparity between mRNA transcripts extracted from yellow and green leaves, but no obvious tissue-specific RNA editing patterns are yet identified among these five genes, although reports in the literature indicated that the extent of atp6 editing is significantly different among tissue types [41]. Therefore we assume the tissue-specific RNA editing patterns may be only detectable in certain genes, cell types, and developmental stages [42].
Plant cells usually possess hundreds to thousands mitochondria or copies of mt genomes that can be regarded as a population when genetic heterogeneity is to be investigated. High throughput next-generation sequence technologies provide us the opportunity to survey single nucleotide polymorphisms in the same or different species (subspecies or cultivars) by mapping reads to a reference sequence and to each cultivar. The polymorphisms within the same cultivar genome (intra-varietal SNPs) and among different cultivars genomes (inter-varietal SNPs), discovered by high-coverage of reads, can also be separated into major and minor genotypes based on simple read counts. Here, we use three runs of SOLiD fragment data from each of the three cultivars (Khalas, Fahal, and Sukry) sequenced in our study for intra-varietal (Table 4) and inter-varietal SNP identification (Table 5). We identified 651, 703, and 731 intra-varietal SNPs in cultivar Khalas, Fahal, and Sukry, respectively, estimated to have a polymorphism rate of one in 1,000 bp, which is about two times higher than that of date palm chloroplast [18] but is only about one tenth of rice mt genome [43]. We should be cautious here since different SNP analysis methods are applied because of the distinct sequencing strategies used in sequencing these mt or cp genomes. The rates of each variation type among these intra-varietal SNPs of the three cultivars are very similar except the types (such as A to T or G to C and vice versa) that do not change GC contents are less represented. These SNPs can also be separated into transition and transversion types, and as a result, there are 297, 325, 287 transitions and 354, 378, 347 transversions for Khalas, Fahal, and Sukry, respectively. The rate of transversions is slightly higher than that of transitions, though in chloroplast transversion (52) is 2× that of transition (26) [18].
All together, there are 120 candidate SNP sites identified among the three cultivars (Table 5), with an inter-varietal polymorphism rate of 0.017%, similar to that of subspecific (between subspecies) polymorphisms between rice cultivar 93-11 and PA64S, ~0.02% [43]. The inter-varietal SNPs are predominantly found in non-coding regions, only seven SNPs were found in coding sequences (all are located in 26S and 18S rRNA genes; Table S7): two between Khalas and Fahal, six between Khalas and Sukry, and six between Fahal and Sukry (Table 5). As to the remaining 113 inter-varietal SNPs residing in non-coding regions (Table S8), 79 of them are between Khalas and Fahal, 91 between Khalas and Sukry, and 50 between Fahal and Sukry (Table 5). Such a distribution implies that Fahal and Sukry are more related than either one of them to Khalas.
P. dactylifera mt genome has much less repetitive sequences as compared to those of other known angiosperms [44]. Only one long palindromic sequence with repeat unit longer than 1000 bp was identified (Table S9) and no inverted repeats were found. Overall, long repeats (>50 bp) only account for 2.3% of the genome, even lower than that of Vitis (2.9%) and Vigna radiata (2.7%) [45], which contain the lowest repeat contents among sequenced angiosperm mt genomes, whereas Tripsacum and Oryza contains 36.4% and 30.4% long repetitive sequences, respectively. This situation also applies to tandem repeats, which constitute only 0.33% of the genome (Table S10). Among the examined 15 plant mt genomes, whose tandem repeat contents range from 0.08% (N. tabacum) to 6.13% (Cycas taitungensis), only three mt genomes, those from N. tabacum, O. sativa and Chara vulgaris, contain tandem repeats lower than date palm (Figure 5). It is well known that plant mitochondria are exceptionally flexible in genome size and structure, and the accumulation of repetitive sequences often results in high sequence divergence. For instance, Cucurbita mt genome contains 38% of short repeats (19–621 bp in length) that make it the largest reported mt genome so far [31], whereas maize expanded its mt genome size by duplication of large sequences [46]. Therefore, it is rather unusual that date palm mitochondrial genome is both lower in tandem repeat content and rare in large duplications. It seems that larger mt genomes of angiosperms tend to have shorter repeat lengths when long repeats are compared. For instance, mt genomes of Cucurbita (982 kb), Vitis (773 kb), and P. dactylifera (715 kb) have their largest repeat lengths of 621 bp, 651 bp, and 1,171 bp, respectively.
The mt-genome is transcribed by a phage-type RNA polymerase encoded in the nuclear genome [47]. The transcription process is rather complex characterized by splicing, editing, terminus processing, and multiple promoters [48]. In addition, mitochondrial genome transcription is reported to be capable of adapting to specific regulation [49]. Here, in order to better understand tissue-specific mt gene regulation and the contribution of mt genes to the development of different tissues, we performed a thorough transcriptome survey across eight date palm tissues (Figure 1 and Figure S2) using high-performance next-generation sequencers. We discovered that ~30.8% regions of our assembly are transcribed (Table 6), slightly lower than that of the rice (~48.5%) [50], with an average sequence coverage of ~44× calculated based on 40 conserved house-keeping genes (Figure 6). On the one hand, our whole genome level gene expression profiling indicated that two tissues, green leaf and fruit, have the most abundant transcripts (Figure S2) but have the lowest gene expression level in terms of RPKM value (reads per kilobase of exon model per million mapped reads) [51] (Figure 6). Male and female flowers, root, and bud, on the other hand, tend to have higher gene expression levels but less in transcript abundance than the leaves. We assume that developing tissues, such as yellow leaf, bud, and root, need more energy than the relatively mature tissues, such as green leaf and fruit. By the same token, the highly expressed genes in female and male flowers are possibly related to flower development that not only depends on a set of nuclear genes but also on the coordinated action of mitochondrial genes [52]. It is possible that the variable expression of mt genome-encoded genes is relevant to the copy number variation of mt genomes (similar to the number of mitochondria per cell) [53], [54] or its changing status in tissue development. In addition, several other obvious tissue-specific gene expression patterns can be observed. First, consistent with a previous study that atp1 gene prefers to express in pollen mother cells [55], we also detected that the transcript of atp1 is obviously more abundant in male flower than in other tissues examined. There is another gene matR that encodes a maturase-related protein also expressed in a relative higher level. Previous study revealed that this gene suffers from modest RNA editing in maize and soybean and was predicted to be functional [56]. Our results here indicate that this gene should have utmost importance in male flower development. Second, the maturation process of yellow to green leaves seems to involve the suppression of about half of the 40 mt house-keeping genes, and the fact is further confirmed that developing tissues are more affected by mt gene expression. Third, during seed maturation, half of the genes were found to be up-regulated when compared to the mt gene expression pattern in fruit. Fourth, interestingly, we found that the gene expression patterns of yellow leaves and seeds are quite distinct–down-regulated genes in one tissue tend to be highly regulated in the other tissue–except that of ccmFn, cox2, rps1, rps3, and rps19 which have no obvious differences between these two tissues. Fifth, two genes, rps1 and rps19, are found clearly highly expressed in root as compared to other tissues. The functional roles they play in root development still need further experimental confirmation. Sixth, consistent with previous studies, rRNA gene transcripts are found to be more abundant, ~9–13 fold than protein coding genes [57], but our large-scale transcriptomic analysis reveals a much higher transcription level changes ~50–400 fold than the average level of protein-coding genes according to RPKM values, and the order of expression levels for the ribosomal RNAs is 5S rRNA >26S rRNA >18S rRNA.
As the first of the palm family plants, P. dactylifera mt genome displays several unique features. First, it positions at the root of the known monocot mt genomes. Second, it has a very low level of repeat content and shows abundant RNA editing events. Third, it exhibits a high level of chloroplast sequence insertions as compared to other known angiosperm mt genomes. Furthermore, our large-scale transcriptome analysis revealed that ~30.8% of its sequences are transcribed and show obvious tissue-specific gene regulation patterns, among which both female and male flowers, root, and bud exhibit higher gene expressions than other sampled tissues. Our complete mt genome sequence assembly represents a new addition to the growing number of plant mt genomes in the public databases and paves a way for further investigations on mitochondrial biology of seed plants.
We used three domestic P. dactylifera cultivars, Khalas (male and female), Sukry (female), and Fahal (male), for this study. Tissue samples from adult date palm trees grown in Al-Hasa Oasis of Kingdom of Saudi Arabia are harvested, including soft bud, flower (male and female), fruit, root, yellow leaf (young), and green leaf (old). We disinfected the samples with 75% ethanol and froze them in liquid nitrogen immediately. For longer term storage, they are stored in −80°C freezer until use.
The P. dactylifera mt genome sequences are produced as part of the Date Palm Genome Project (DPGP, a joint effort between KACST and CAS). Genomic DNA was extracted from 50 g soft bud tissues according to the CTAB-based method. We used 5 µg purified DNA for shearing and constructing fragment libraries following the GS FLX Titanium general library preparation protocol. The ssDNA libraries were amplified with emulsion-PCR and enriched, and the samples were sequenced on Roche/454 GS FLX platform.
SOLiD long mate pair (LMP) libraries of the three cultivars were constructed by following SOLiD Library Preparation Guide (SOLiD 4.0) and at least 20 µg genomic DNA was used depending on different insert sizes (600–6000 bp). After emulsion PCR and beads enrichment (EZ beads system, AB), template beads of each LMP library were deposited to 2 quarter of slide and then loaded onto a SOLiD 4.0 instrument.
For transcriptomic study, tissue samples were grinded into fine powder followed by CTAB-based RNA extraction, and 2.5 M LiCl was used to remove polysaccharides. 0.5 µg rRNA-depleted total RNA (RiboMinus Plant Kit, Invitrogen) were used to construct transcriptomic libraries according to the instruction from SOLiD Total RNA-Seq Kit.
We separated candidate mt genome reads from eight Roche/454 GS FLX runs based on 40 published plant mt genome sequences (identity ≥80% and E-value ≤10−5). About 1.5 millions reads were obtained and assembled by using Newbler (version 2.3 with default parameters)–a de novo sequence assembly software provided by Roche. As a result, we obtained 29 mt genome contigs (total ~438 kb) with an average length of 15 kb. These contigs were extended to 662 kb by adding additional Roche/454 reads. Subsequently, SOLiD mate-pair data (2×50 bp libraries) with insertion sizes of 1–2 kb and 3–4 kb were used to construct scaffold (50-nt overlap cutoff and less than 2-nt mismatch). A total of 3,918 homopolymers with repeat unit ranging from 5 to 11 were verified and revised based on SOLiD fragment data using BFAST program (version 0.6.4d) [58]. At last, 715,001 bp complete mt genome was assembled with an average sequence depth 130×. The final genome sequence was validated by SOLiD LMP data with insertion sizes of 3–4 kb, 4–5 kb, and 5–6 kb in a 2 kb sliding window with variable step sizes; we show the result from an analysis using 5–6 kb insert size in 15-kb step size in Figure 1.
The complete sequence of the date palm mt genome was deposited to GenBank (accession number JN375330).
A preliminary annotation was carried out by mapping final genome sequence with BLAST (identity >90% and overlap>90%) [59], [60] hits to known mitochondrial genes, and subsequently by testing for consistency of the ORFs using NCBI online tool the ORF finder (http://www.ncbi.nlm.nih.gov/projects/gorf/, the standard genetic code was applied). The exact gene and exon boundaries were determined by alignment of homologous genes from several common mt genomes (Table S2) and verified based on transcriptomic data. The tRNA genes were identified by using a local chloroplast and mitochondrial tRNA database, BLAST search tools [59], [60], and the help of tRNAscan-SE program (version 1.4 and default parameters were used) [61]. Both group I and group II introns were predicted by using an online software Rfam (version 10.1; http://rfam.sanger.ac.uk/; default parameters) [62]. Homology search using BLAST [59], [60] was carried out to identify chloroplast-derived regions in the mt genome assembly (over 80% sequence identity; E value ≤1e-5; length >50 bp).
We predicted putative RNA editing sites in protein-coding genes using the PREP-mt web-based program (http://prep.unl.edu/) [40]. To achieve a balanced tradeoff between the number of false positive and false negative sites, the cutoff score (C-value) was set to 0.6 as suggested by the author. All other parameters are set to default values. We also verified some of the RNA editing sites in five genes (atp1, atp4, atp9, rpl116, and rps19) across the two leaf tissues (yellow and green leaves) using cDNA data from Roche/454 GS FLX system (NCBI accession number SRA045434.3). The five genes are chosen randomly, whose cDNA sequences are full-length and better in quality.
We carried out both intra-varietal and inter-varietal SNP analysis across three cultivars: Khalas, Fahal, and Sukry. Three runs of SOLiD LMP reads for each cultivar (about 60 Gb) were mapped to the reference mt genome (Khalas) by using BioScope software (version 1.3). The mapping results were then used for SNP identification based on a Bayesian algorithm according to the BioScope Software User Guide.
We identified repetitive sequences, including forward, palindromic, reverse, and complemented repeats, using the REPuter (version 2.74; with a minimal length of 50 bp and 3 mismatches) [63]. We removed overlapped repeats manually and obtained information on tandem repeats using a tandem repeat finder (http://tandem.bu.edu/trf/trf.html; default parameters were used) [64].
We used 22 protein-coding genes (atp1, atp4, atp6, atp8, atp9, ccmC, ccmFn, cob, cox1, cox2, cox3, nad1, nad2, nad3, nad4, nad4L, nad5, nad6, nad9, rps3, rps4, and rps12) common to 15 plant mt genomes for our phylogenetic analysis. We aligned the sequences using Clustalw2 (default parameters were used) [65], removed ambiguously aligned regions based on Gblocks (version 0.91b; minimum number of sequences for a conserved position and flanking position is set to 10 and 15, respectively; no more than eight contiguous non-conserved positions and no gap are allowed) [66], and concatenated the sequences. The maximum-likelihood tree was constructed by using PHYML (version 3.0) [67] under HKY85+Γ4 model (C. vulgaris is used as outgroup). The bootstrap value was set to 100. All other parameters are set as default.
We used transcriptome data from bud, root, seed, fruit, male and female flowers, yellow and green leaves of cultivar Khalas. On average, ~700,000 SOLiD reads (50 bp with 3 mismatches or less) are used from the libraries. RPKM values are measured (reads per kilobase of exon model per million mapped reads) [51] and used to estimate gene expression, which are calculated according to:
GC content variations between new and old chloroplast-derived sequences. We defined chloroplast-derived sequences unique to P. dactylifera mitochondrial genome as "New" and those shared by other plant mt genomes as "Old".
Transcriptome analysis across eight tissues. FF, female flower (~422,000 reads); MF, male flower (~589,000 reads); F, fruit (~1,048,000 reads); S, seed (~179,000 reads); B, bud (~457,000 reads); GL: green leaf (~2,388,000 reads); YL, yellow leaf (~606,000 reads); R, root (~545,000 reads); P, genes on the positive strand; N, genes on the negative stand; and CP, chloroplast-derived regions. Their RPKM values (transformed using log10) range from 0 to 9 for genes on the positive strand and 0 to −9 for genes on the negative strand.
15 plant mt genomes used in this study.
Genes in 12 angiosperm mt genomes.
tRNA gene content of P. dactylifera mt genome.
Chloroplast-derived sequences in P. dactylifera mt genome.
RNA editing sites in three different plant mt genomes.
RNA editing validation of five genes in two tissues based on GS FLX reads.
Inter-varietal SNPs in coding regions among the three cultivars.
Inter-varietal SNPs in non-coding regions among the three cultivars.
Long repeats (repeat unit >50 bp) in P. dactylifera mt genome.
Tandem repeats in P. dactylifera mt genome.
We thank Jiucheng Liu, Dawei Huang, Guiming Liu, Quanzheng Yun, Sun Zhang, Guangyu Zhang, Duojun Zhao, Jixiang Wang, and Gaoyuan Sun for their excellent technical supports and contributions to this project. We also thank Professor Jun Tan for his helpful comments and discussions.
Competing Interests: The authors have declared that no competing interests exist.
Funding: The project is supported by a project grant from King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia [428-29]. The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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