Horton, Oak Ridge National Laboratory;and former LBNL Operations DeputyDirector Klaus Berkner. The committeewas staffed by Dan Drell and KentLohman of DOE Office of Biological andEnvironmental Research. JanePeterson, the Associate Director fromthe Division of Extramural Research atthe National Human Genome ResearchInstitute of the National Institutes ofHealth, sat in on the proceedings as anobserver. The areas covered in thereview included both science and man-agement and operations. A verbal sum-mary of the report was delivered by MelSimon to BERAC at the beginning ofDecember and was generally very posi-tive, especially about DOE JGI science.

The U.S. Departments of Agricultureand Energy have announced that theywill share resources and coordinate thestudy of plant and microbial genomics,and DOE JGI will tackle the sequencingof the soybean genome as the first proj-ect resulting from the agreement.

“This agreement demonstrates ajoint commitment to sup-port high-qualitygenomics research andintegrated projects tomeet the nation’s agricul-ture and energy chal-lenges,” said Dr. ColienHefferan, administrator ofUSDA’s Cooperative State Research,

Extension and Economics Service(CSREES), who signed the agreementfor USDA.

Soybean, or Glycine max, the world’smost valuable legume crop, is of partic-ular interest to DOE because it is theprincipal source of biodiesel, a renew-able, alternative fuel. Biodiesel has thehighest energy content of any alterna-

tive fuel and is significantly more envi-ronmentally friendly than

comparable petrole-um-based fuels,

since it degradesrapidly in the environ-

ment. It also burnsmore cleanly than conven-

tional fuels, releasing only half of thepollutants and reducing the productionof carcinogenic compounds by morethan 80 percent. Over 3.1 billionbushels of soybeans were grown in theU.S. on nearly 75 million acres in2004, with an estimated annual valueexceeding $17 billion, second only tocorn and approximately twice that ofwheat. The soybean genome is about1.1 billion base pairs in size, less thanhalf the size of the maize or human

On November 16 through 18, 2005,a panel of eminent scientists accompa-nied by research administrators of theDepartment of Energy’s Biological andEnvironmental Research AdvisoryCommittee (BERAC) met at the DOE JGIPGF to conduct a comprehensive reviewof the DOE JGI. Chaired by Mel Simonfrom the California Institute ofTechnology, the committee includedBruce Birren, the Broad Institute; JaneRogers, The Sanger Centre; RichardWilson, the Genome Sequencing Centerat Washington University St. Louis; JimTiedje, Center for Microbial Ecology atMichigan State University; RichardMural, Windber Research Institute;Bruce Chrisman, Fermilab; Linda

inside this issue2. Spotlight on ORNL/LSP

3. Spotlight on LANL

4. DOE JGI Faces—Tijana Glavina del Rio

6. Spotlight on Safety

7. Critter in the Queue—Trichoplax

8. Tech Transfer

10. René’s Googling Tips

PRIMERthe

USDA and DOE to Coordinate Researchof Plant and Microbial Genomics

Spring 2006 Volume 3 Issue 1

cont. on page 8

cont. on page 8

DOE JGI Reviewed by BERAC Subcommittee

Soybean to Be Sequenced

Save the DateFirstAnnualDOE JointGenomeInstituteUserMeeting

March 29-April 1http://www.jgi.doe.gov/usermtg/index.html

annotation process is guided by the prin-ciple that, if genes found in two differentorganisms look similar, there is a highprobability that they will function in asimilar way.

Loren Hauser, of Oak Ridge NationalLaboratory, explains: “If the sequencedDNA contains a gene that looks likeanother gene which has been experi-mentally proven to be responsible for

the production of a specific protein,there is a high probability that our genehas the same function.”

Two types of organisms are sequen-ced by DOE JGI: microbes and eukaryotes.The two groups differ dramatically, andare handled at DOE JGI through two dis-tinct pipelines. Igor Grigoriev leads theeukaryote annotation effort at the DOEJGI Production Genomics Facility (PGF).Microbes, meanwhile, are sent out forautomated annotation to Hauser andMiriam Land at Oak Ridge. A look at themicrobial pipeline illustrates howautomation is helping to speed up theannotation process.

The microbial annotation pipelinebegins with Sam Pitluck at the PGFuploading data from completely assem-bled microbial genomes to the DOE JGIFTP website, where Miriam and Lorenpick it up. Then, with the help of such

Spring 2006 Vol. 3, Issue 1

2 / THE PRIMER

grams. Second, it willprovide small-scalesequencing that meetsthe needs of individualinvestigators at thenational laboratories.The LSP is expected touse 15 to 20 percent ofDOE JGI’s sequencingcapacity, which is cur-rently over 35 billionbases per year.

Sequencing to becarried out under the LSP will includegenomes of entire microbial communi-ties, and individual microbes andplants, useful in decreasing reliance onpetroleum and petrochemicals by con-

verting plant materials, such as soy-beans, to ‘green’ energy and chemicalfeedstocks. Sequencing will also focuson characterizing the variation in humansusceptibility to nucleic acid damage byionizing radiation.

LSP’s newly appointed lead isGerald Tuskan, a senior scientist in theEnviron-mental Sciences Division at OakRidge National Laboratory. Tuskan willbe responsible for developing, coordi-nating, and managing the LSP. Tuskangot together with DOE JGI’s DanielRokhsar and colleagues from theNational Center for SoybeanBiotechnology, the USDA AgriculturalResearch Service, Purdue University,and the National Center for Genome

As hundreds of genomes churnthrough DOE JGI sequencers, efforts areunderway to make their annotation—thedetermination of what biological functionsthese sequences actually encode—faster,more accurate, and cheaper. One of thechallenges in assigning function tosequence is that the generation of rawsequence has far outpaced experimentaldata-gathering. To help close this gap, the

SPOTLIGHT ON DOE JGI ORNL

MICROBIAL GENOME ANNOTATION THE ORNL WAY

The LaboratoryScience Program (LSP) isa new initiative from DOEto leverage JGI sequenc-ing capacity, providingDOE national laboratoryresearchers with broaderaccess to high-throughputDNA sequencing in sup-port of mission-relevantprojects.

The LSP will servethe national laboratoriesin two major ways. First, it will fosterlarge-scale strategic sequencing proj-ects, across the national laboratory sys-tem, that are aligned with future fund-ing opportunities in DOE’s biology pro-

ORNL’s Tuskan to Lead LSPcont. on page 12

cont. on page 14

Finished GenomePGF/LANL/LLNL

Automated Annotation

ORNL

Annotation QCPGF

GenBankSubmission Prep

ORNL

GenBank SubmissionPGF

IMG/PortalSubmission

PGF

DOE JGI Microbial Genome Annotation Pipeline

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 3

BY KENDRA MYERS, LANL STUDENT

Introduced in December 2004, DOEJGI LANL’s ongoing Genomic ExplorersSeminar Series features researchersfrom various institutions who are collab-orating with the DOE JGI on the genomicsequencing of a particular organism.

The LANL community looks forwardto the monthly seminars, which giveresearchers and students a chance tolearn about microbes that have beenselected and prioritized, through a highlycompetitive review process, forsequencing at the DOE JGI. The semi-nars also provide an opportunity for thespeakers to learn about DOE JGI LANL’scapabilities in genome sequence finish-ing, and annotation and analysis.Speakers typically spend a day or two

in Los Alamos, enough time to meetwith scientists who may have con-tributed to their sequencing project andwith other researchers who may havesimilar interests.

For the most part, speakers invitedfor the seminar series are part of eitherthe Community Sequencing Program(CSP) or the Microbial Genome Project(MGP). By welcoming submissions fromresearchers outside the DOE JGI forgenomic sequencing of an organism,these programs make the large-scalesequencing capabilities of the PGF avail-able to the greater scientific community.Organisms are selected based on theirpotential contributions to the DOE mis-sions of advancing bioremediation, car-bon sequestration, alternative energydevelopment, and biotechnology, to

name a few. About half of the seminarsat LANL focus on bacterial organismsand the other half on fungal organisms.

In February, Teresa Thiel, from theUniversity of Missouri, gave a seminaron a cyanobacterium that was selectedfor sequencing based on its ability to fixN2 and CO2, and to produce H2. This

BY KATHERINE HARRINGTON AND REBECCA E.

MCINTOSH

Plants are essential commoditiesfor many industries, from food to biofu-el, textiles, and building materials.Scientists at Los Alamos NationalLaboratory (LANL) are protecting ournation’s agricultural investments usingmethods originally designed to identifyhuman pathogens. The Plant PathogenConsortium was formed a little over ayear ago to tackle the problem ofpathogens that threaten major crops—pathogens that, because of the eco-nomic importance of these crops, are insome cases considered potential bio-threat agents. The Consortium’s efforts

focus on developing the early detectiontechniques and effective treatmentsessential for the quick response need-ed to prevent major agricultural losses.

Lacking the ability to produce anti-bodies or other adaptive responses topathogens, plants have limited defens-es against disease. Therefore, the prior-ity for farmers is to prevent plants frombecoming sick in the first place. Whenthis fails, there is a need for earlydetection and for non-toxic treatmentssafe enough to use alongside healthyplants. Naturally, scientists would liketo identify the pathogen when it hasaffected only a few plants instead ofthe entire crop, thus preventing the eco-nomic hardship of losing an entire har-

BY REBECCA E. MCINTOSH

After six years at Walnut Creek,Chris Detter has decided to make amove to greener, nope, browner pas-tures. Chris, who led the cloning tech-nology group at the DOE JGI PGF,recently accepted a position at DOE JGILANL and left sunny California for north-ern New Mexico.

New Mexico isn’t a completely newplace for Chris. Over the past fouryears, Chris has collaborated withresearchers at LANL to develop moreefficient DNA cloning protocols for theBioscience Division. In his new posi-tion, Chris will contribute to the finish-

DOE JGI Assists in Plant PathogenConsortium Research

Detter Joins Teamat DOE JGI LANL

Genomic Explorers Seminar Series

cont. on page 14

cont. on page 11

cont. on page 15

SPOTLIGHT ON DOE JGI LANL

Teresa Thiel, Professor of Biology,Associate Dean, College of Arts &Sciences, University of Missouri, St. Louis.

Spring 2006 Vol. 3, Issue 1

4 / THE PRIMER

ral gas company. “All his life he’s beeninvolved in management. I get a lot ofmy management skills from him, partic-ularly the ability to work with people,understand where they are coming fromand coach them to ensure success.”

Naturally, Tijana is a believer in thepower of genes. “I look at my parentsand see what kind of traits I’ve gainedfrom each of them.”

While she tried to concentrate onher studies, the war rumbled on. “A lotof young boys went to the war and gotkilled. There were times when the actiongot close to downtown. One day a bombfell—it was one of those grenades thatsplit into many pieces—right inside thisrehearsal room. It had a glass ceilingso people got hurt. Girls I know gotshrapnel all over their feet. Can youimagine, for dancers!”

The event was sufficient motivation tomake the move to California. She hadrelatives in the Golden State and came fora visit in 1990. “I liked it a lot in WalnutCreek and San Francisco. I mean, what’snot to like? I decided I was in love withCalifornia and one day I was going back.”

The part of the state where Tijanaended up moving to was far from the

Tijana Glavina del Rio has been partof the DOE JGI Production GenomicsFacility (PGF) since shortly after the doorsopened back in 1999, but Tijana’s pathto the PGF was anything but conventional.

Growing up in Croatia, she was inthe water practically before she couldwalk. “I got a BS with a concentrationin marine biology because I was hopingto go work with the whales and dolphins,explore the oceans. I was really in lovewith Jacques Cousteau and his explo-rations since I was a little kid.

“However, when I graduated, therewere just too many marine biologistsand not enough jobs, and a BS didn’tprepare me enough for those jobs. So Istarted looking for anything that had todo with biology. I stumbled upon theHuman Genome Project, which caughtmy attention. Then I saw an applicationfor a lab tech position at LawrenceLivermore National Laboratory.”

She never actually worked inLivermore, though, because she wasneeded straightaway in Walnut Creek.“Nancy Hammon was my coach duringthe first couple of weeks at JGI. Shetrained me on cleaning up DNA samplesusing the SPRI [Solid Phase ReversibleImmobilization] original bead cleanupmethod. At the time, we were just form-ing a production line. Our throughputwas just one 96-well plate per day.Next, they brought the CRS robots and Iwas tasked to work on automating theSPRI process using the new robots.”

Tijana’s responsibilities have grownby leaps and bounds over the last fiveyears, and today she is the SequencingPrep Group Leader. She now supervis-es, in addition to sequencing prep, theFreezer Group and the Fosmid Group.It’s quite a complicated dance, butshe’s up to the task.

From an early age Tijana, an onlychild, attended a classical ballet school

in the Croatian capi-tal city of Zagreb.“Zagreb’s a beautifulcity. It was part ofthe Austro-HungarianEmpire, so it hasvery similar architec-ture to Vienna andPrague. The whole ofCroatia is breath-taking.

“I started attend-ing classical balletat the age of fiveand was quite seri-ous. With the balletschools over there,you either sink or swim. They are verycompetitive. My mom kept me in therebecause I was doing well. I graduatedfrom classical ballet school and earneda diploma at the same time as my highschool diploma. My school schedulewas much regimented; I never had anyfree time. After schooling, I startedworking in the National Theater ofCroatia, where I danced as part of anensemble for three years. I played rolesin many beautiful ballets, from DonQuixote to Swan Lake, the Nutcracker—anything you could imagine.

“When I enrolled at the University ofNatural Sciences in Zagreb as a fresh-man it was very difficult and challengingto keep up with schoolwork and dancing,so eventually I had to make a careerdecision. I chose biology, but never lostthe love for ballet. We call it ‘bloodywork,’ because your feet bleed. Youwork so hard and they pay you so miser-ably. But, nevertheless, I really enjoyedevery minute of it.”

Tijana drew her inspiration andtenacity from her parents. Her mother isa professor of English and French, andher father is a mechanical engineer andmain production manager of a big natu-

cont. on page 5

PGF Sequencing Prep Group Leader Tijana Glavina del Rio

JGI FACES

Tijana Glavina del Rio—Biomedical Ballerina

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 5

A Berkeley Lab Outstanding Perfor-mance Award (OPA) has been presentedto a team, consisting of Dana Alcivare,Jason Baumohl, Patrick Hajek, AaronPorter, and Arkady Voloshin for their cre-ative and exceptional accomplishment inporting the legacy Venonat LIMS from aproprietary, obsolete, unsupported plat-form to an industry-standard Oracle 10g

database and Apache web server.Common wisdom held that it was notpossible to upgrade WebDB applications;the team was confronted with a 10+person-year effort to rewrite the wholeapplication. The porting breakthroughand implementation saved a huge amountof time and effort that was better spenton improving the application.

New Safety Officer StephenFranaszek has a Bachelors in Mech-anical Engineering and a Masters inEnvironmental Health Science fromUniversity of California at Berkeley.He has 15 years’ experience in Safetyand Industrial Hygiene at LLNL andon the UC Berkeley Campus. Forthe last three years he has beenLLNL Engineering Safety Officer.

In October2005, Ray Turnerassumed the helmof the DOE JGIOperations Depart-ment, where heoversees finance,human resources,facilities, and

administration at the PGF. Ray receivedhis BS in Finance from the University ofUtah and was commissioned into theUnited States Navy in 1980. After com-pletion of flight training he served invarious senior management and trainingpositions within the Navy. In 1995, hereceived a Masters Degree in FinancialManagement from the Naval Post Grad-uate School and served as the Comp-troller at Naval Air Station Alameda dur-ing the base closure process. Aftercompletion of a 21-year Navy career, hejoined a San Francisco Bay Area health-care information technology companywhere he served as the Vice Presidentof Finance and Vice President/GeneralManager of the Health InformationManagement Division.

NEW OPS HEAD RAY TURNER

JGI New Faces (Sept 2005–March 2006)PAUL BARALESoftware Developer 3, Instrumentation

MATT BLOWComputational Biologist PostdocFellow, Vertebrate Program

BILL CANNANSr. Recruiter, Human Resources

ARYK GROSZSoftware Developer 1, Instrumentation

ELIZABETH (BETTE) HERRERAOperations Administrator, Operations

NATALIA MIKHAILOVASystems Analyst 2Genome Biology Group

ROBERT OTILLARSystems Analyst 3, Genome Annotation

MICHAEL PHILIPSBiosciences Technician, Production

ROTEM SOREKComputational Sciences PostdocFellow, Vertebrate Program

ALICIA TOLIBASHR Asst III, Human Resources

RAY TURNEROps Dept Head, Operations

PARAG VAISHAMPAYANComputational Biologist PostdocFellow, Vertebrate Program

OPA FOR PORTING VENONAT

Tijana—Biomed Ballerina

beach and the whales: on a campus,Cal State Stanislaus in Turlock, that’slandlocked in California’s great CentralValley. The school is known by locals as“Turkey Tech,” because of Turlock’s rep-utation as a leading poultry producer.

“I had no idea where I was goingbut I was so desperate to leave Croatiathat it did not matter. When I got there,I couldn’t smell anything but cows. Icried the first night. I was thinking that Iwanted to go back home. The new envi-

ronment was a shock. Where’s the beach?Where is all the beautiful stuff I’d seenon my first visit? I had no friends. It tookme some time to adjust. But luckily forme, it only took two days to meet my soulmate.” Her husband Daniel, originally fromSpain, spent most of his life in Tenerife,in the Canary Islands. Ten years laterthey have two American kids, Nicolasand Isabel, who are destined to be per-fectly trilingual in Spanish, English, andCroatian. But will they dance?

cont. from page 4

Spring 2006 Vol. 3, Issue 1

6 / THE PRIMER

To increase awareness of ergonomicsas part of the safety culture at the PGF,production groups will be putting togetherErgonomics posters for their workingareas in the production facility. Look forposters in each of the labs as each staffmember has identified ergonomic risks,ergonomic changes, and possible solu-tions to help keep their work area safeand efficient. Everyone is gettinginvolved with the goal to keep our facilitya safe place to work.

FOCUS ONERGONOMICSKeeping the PGF on the SafetyTrack

Implemented last December, SafetyTrack is the Production GenomicsFacility’s centralized safety issue tracking system. Through SafetyTrack, you

can report a safety incident or register a safety concern which could result inan incident if not addressed. You can also anonymously track the status of

anonymous submissions.

Check out http://venonat.jgi-psf.org/safetyTrack to:• Report incidents • File requests • Voice concerns •

• Leave feedback • Audit concerns •

SPOTLIGHT ON SAFETY

“We have taken a major stepwith these internal appointmentsto implement a structure thatallows us to offer internal leader-ship opportunities, increase pro-duction capacity, and provide thenecessary support structure tooperate a production staff sevendays a week, two shifts in a safework environment,” said SusanLucas, DOE JGI SequencingDepartment Head.

PROMOTIONS—NEW PRODUCTION SHIFT LEADS

A show of hands for SafetyTrack by Tijana Glavina del Rio, Chris Hack,and Terri Jackson.

Amber Nivens Matt Zane Eric Abbott Cailyn Spurell

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 7

BY WENDY SCHACKWITZ

Welcome to the first editionof the “Critter in the Queue” col-umn. The goal of the column isto explore the “what” and “why”behind the organisms the DOEJGI is sequencing. It is an oppor-tunity to turn the cryptic namesscribbled on the side of asequencing plate into real livingcreatures, and to understand whythe DOE JGI has chosen tosequence their genomes. Thefirst Critter in the Queue isTrichoplax adherens.

This is a Community SequencingProgram (CSP) project that is beingcompleted in collaboration with StephenL. DellaPorta (proposer) and Leo Buss,Yale University; Dan Rokhsar (propos-er), DOE JGI; Bernd Schierwater,Tieraerztliche Hochschule, Germany;Rob DeSalle, American Museum ofNatural History; Peter Holland,University of Oxford; and GeorgeWeinstock, Baylor College of Medicine.

Trichoplax Basics

At first glance, this critter looks alot like an amoeba, but it is very differ-ent. While an amoeba is a single cell,Trichoplax is a true animal, albeit a verysimple animal.

At only a few millimeters in diame-ter, it has the simplest body plan of anyanimal, consisting of only four celltypes that are arranged in only threecell layers. Based on its appearance,zoologist Franz Eilhard Schulze in 1883named this animal Tricoplax adherens,which in Greek means sticky-hairy-plate.

Trichoplax is found in tropical andsubtropical oceans around the world,where researchers have been able tocollect these animals from the water

column. They have also been found liv-ing on a variety of subtidal surfaces,including on other animals such as fish.Because studying Trichoplax in its naturalenvironment has not yet been possible,very little is known about it.

Some cool facts:Unlike most other animals it lacks a

basal lamina, a layer of extra-cellularmaterial that is used to organize tissuesin most other animals. Some groups ofsponges also lack a basal lamina.

Unlike all other animals it seems tolack an extracellular matrix, the materialthat exists between cells that assists incell-to-cell adhesion and communication.

Why is DOE JGI sequencing it?

PHYLOGENETICS Trichoplax is so unusual that it has

been placed in its own phylum, thePlacozoa. Furthermore, it has beenhypothesized that this animal divergedfrom other animals soon after multicel-lularity evolved. Although its exact posi-tion in the tree of life remains uncer-tain, it seems clear that Trichoplax rep-resents an early branch of the animaltree of life. Therefore, understandingsimilarities and differences betweenTrichoplax and other animal lineages will

give us unique insight into animalevolution. It will also provide anew standard basal genome forcomparative analysis.

Minimal Genome

Trichoplax’s genome is 50megabases, the smallest of anyanimal. This is only ten times larg-er than the genome of the bacteri-um E. coli and three times largerthan yeast.

Cancer and Aging

Trichoplax has only beenobserved to reproduce asexually,

either by dividing in half or by buddingof small “babies” called swarmers.There is evidence that this animal hadthe ability to reproduce sexually in itsrecent evolutionary past, but whether ornot it retains this ability presentlyremains uncertain.

Trichoplax is the first animal to haveits genome sequenced that can repro-duce asexually. Some of its laboratorycultures have been maintained continu-ously via asexual reproduction for morethan 20 years. Without sexual reproduc-tion, which provides special DNA repairmechanisms, it would be expected thatthis culture would suffer from the sameproblems our bodies experience withtime that lead to aging and cancer, yetthis is not seen in these cultures. Sohow do they keep from aging or gettingcancer? Do they have special DNArepair mechanisms or other ways to pro-tect themselves from oxidative damageand telomere degradation? Once thegenome is sequenced, we will be betterable to answer these questions.

How Far Have We Gotten?

The Trichoplax genome draft isexpected to be completed this summer.

CRITTERS IN THE QUEUE

Spring 2006 Vol. 3, Issue 1

8 / THE PRIMER

Among the highlights cited by Simonon the BERAC website were:

• The experts were quite satisfied.• Appropriate and inspiring vision. • Driving and exploiting sequence-

based science. • Have “cornered the market” on

sequencing for energy, carbonsequestration, and environmentalremediation.

• DOE JGI line managers are very good,especially the sequencing lead.

The written draft report now goes tothe full BERAC committee, which isexpected to convene in May for reviewand (probable) approval of the report.

The Technology Transfer group atthe PGF, supervised by Nancy Hammon,facilitates the planning, coordination,and implementation of new technolo-gies, protocols, and workflow into theproduction sequencing line while contin-uously communicating the status ofthese transitions effectively acrossmany DOE JGI groups. The TechnologyTransfer group is also responsible forproduction line project implementation.Projects include any new instruments,protocols, reagents, hardware, software,or workflows. Project responsibilitiesbegin at the earliest formation of anidea and extend through the productionadoption, but are primarily concernedwith a project’s transition from workingmodel to production line ready.

Current projects include:

FOSMID 384 PROTOCOL (Jamie Jett,Diana Lawrence, Julita Madejska,Maryam Waheed, and Duane Kubischta)

A new fosmid protocol has beendeveloped and implemented in produc-tion. This protocol introduces a 384-wellplate format for the fosmids, which willaccount for a 30 percent fosmidreagents and consumables cost sav-ings. The new protocol also gives us atheoretical maximum throughput of 64fosmid plates per day, over twice thecurrent maximum. We are currently run-ning Biomeks 5 and 6 with the fosmid384 protocol. A new version of theSprint Prep reagents is being testedside-by-side with the current version ofSprint Prep. The two versions will con-tinue to run in parallel as we refine,evaluate, and optimize the workflow.

1/32ND BIG DYE CHEMISTRY (DavidHillman, David Robinson, and Mingkun Li)

In December 2005, a coordinatedeffort lead by the Technology Transfergroup set out to implement the newestversion of the 1/32 BigDye sequencing

reaction handed down from GenomicTechnologies. Based on a statisticalanalysis performed by Mingkun Li, the1/32nd BigDye samples have at most aloss of 24 Q20 bases. The library beingsequenced and the polymer lot on thesequencer are significant contributors tothis loss in quality. With the consent ofthe Sequencing Preparation Supervisorand QC, Technology Transfer launchedthe full-scale production use of the1/32 BigDye chemistry reaction onJanuary 25.

NEW PCR PLATE FOR PRODUCTIONSEQUENCING (Victor Hepa, DaveRobinson, Nora Nichols, Sharon Ropes,and Simon Roberts)

Several new PCR plate types havebeen brought in for evaluation in orderto address an ergonomic concern ofplates sticking in PE Thermocyclers andwarping of the PCR plate due to heat.These plates have been the Axygen M2plate, the Eppendorf twin.tec plate, andthe Axygen Hard Shell plate. After someinitial evaluations, the M2 plate was notfound to significantly reduce ergonomicstress on the operators and sufferedfrom the same warpage as production’scurrent Axygen plate. The Eppendorftwin.tec plate is still under investiga-tion. So far, this plate shows promise inboth reducing ergonomic stress andplate warpage. Negotiations are ongo-ing to make this plate cost competitive.The Axygen Hard Shell plate failed itsinitial testing phase due to severe platewarping. The vendor is currentlyaddressing this issue. Some changesto workflow and instrument heightshave been made in the lab to reducethe overall ergonomic concern for thistask while the new plates are underinvestigation.

genomes. “The soybean represents an excel-

lent example of how DOE JGI is playinga key role in ‘translational genomics,’that is, applying the tools of DNAsequencing and molecular biology tocontributing to the development of newavenues for clean energy generationand for crop improvement,” said DOEJGI Director Dr. Eddy Rubin. “Effectiveapplication of translational genomics tosoybean requires detailed knowledge ofthe plant’s genetic code. With this start-ing material in hand, researchers inacademia, industry, and agriculture willbe better positioned to optimize soy-bean for the broadest range of uses.”

Transferring Technology at the PGF Every Moment Spent Planning Saves Three or Four in Execution

Soybeans BERACcont. from page 1 cont. from page 1

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 9

BY CAILYN SPURRELL

Thousands of students, families,and teachers gathered at OrindaIntermediate School on October 6,2005, to extract DNA, gaze at thestars, use liquid nitrogen to make icecream, observe farm animals, and joinin many other activities. Designed touse “way cool science stuff” to raiseawareness among middle schoolers ofscience in general and careers in sci-ence in particular, the event combinedclassroom demonstrations, outdoorbooths, and large presentations.Teachers, community members, andscientists from Bay Area companies,the University of California, StanfordUniversity, LLNL, and LBNL captivatedattendees with their enthusiasm for sci-ence and the lively topics covered in atotal of 36 inventive demonstrations.The atmosphere was electric, as kidsswept through the carnival of scientificexperiments and presentations, pullingalong parents and friends from one dis-play to the next.

Volunteers from the DOE JGI, led byDavid Gilbert, worked with genome edu-cation outreach coordinator Karen Kellyof Clayton Valley High School to engagestudents of all ages in a hands-onexperiment to extract DNA from blue-berries, strawberries, and raspberries.A legion of Miramonte High School vol-unteers helped prospective scientistsinto lab coats, gloves, and goggles, andDOE JGI volunteers Damon Tighe, JulitaMadejska, and Cailyn Spurrell instructedeach table of four or five participants.

The berries were first mashed usinga mortar and pestle. They were thencombined with an extraction buffer con-sisting of meat tenderizer, detergent,and salt before being gently mashedsome more. The reaction mixture wasseparated from the seeds and pulp of

the berries using cheesecloth. Ethanolwas slowly added on top of the filtrate.Instantaneously, small bubbles formedin the alcohol layer, and eyes shone asthin white strings of DNA precipitated.Students were able to collect the DNAusing an unfolded paperclip and take ithome in a small vial of ethanol.

A continuous stream of children andparents were eager to try their hand atrecovering DNA from a small berry, andwere intrigued when a comparativelylarge stringy mass precipitated in theethanol. Orinda Intermediate students,with their younger siblings and friends

from nearby schools (and a few parents),tried the protocol. Some mashed withvigor, messily combining the fruit extractand ethanol or violently shaking up themixture. However, the experiment wasextremely forgiving, and all reactionsproduced some amount of DNA.

After the extraction was performed,DOE JGI volunteers engaged the stu-dents in short discussions about theextraction to illustrate that the samemolecule of DNA is present in all livingorganisms and that it is a real, tangiblething. Each group also learned somequick cell biology, discussing what eachstep of the reaction accomplished inthe cell and how the DNA was extracted.Seeing the excitement and awe on thestudents’ faces as they became scien-tists, extracting berry DNA, was anenjoyable and valuable experience. Ifyou would like to participate in futureevents, contact Karen Kelly(kkelly@lbl.gov).

DOE JGI Delivers “Way Cool” DNA Activity

PGF's Julita Madejska doles out the pipettingpointers to the next generation genomicistswhile five-year-old Alyssa Tsuyuki (below)marvels at strawberry DNA at the OrindaIntermediate School Science Fair.

Spring 2006 Vol. 3, Issue 1

10 / THE PRIMER

ples for searching the DOE JGI intranet:• Search for Microsoft Word docu-

ments containing the word“hybrid” (filetype:doc hybrid).

• Search for web pages that havethe word “ramorum” in their title(filetype:html intitle:ramorum).

• Search for Microsoft Word docu-ments on the DOE JGI wiki thathave the word SOP in their title(filetype:doc inurl:wiki intitle:SOP).

Note: the title of a word document is notthe printed title, but the title in the docu-ment properties.

• Search for all the pages that ref-erence (link to) theChlamydomonas reinhardtiigenome portal page (link:http://genome.jgi-psf.org/chlre2/chlre2.home.html).

Note: the link operator should be usedalone. No other operators or keywordsshould be part of the search string.

• Search for all non-html and non-pdf documents containing theword “Halloween” located some-where else besides the DOE JGIwiki (Halloween -inurl:wiki -file-type:html -filetype:pdf).

Other Considerations

ALTERNATIVE SPELLING(S)If your query didn’t return any

results (or not the results you wereexpecting), try searching again with adifferent spelling (and the OR operator).For instance, if searching for “eRoom”didn’t find the document you were look-ing for, try “eRooms” (or “eRoom OReRooms”). Note 1: the www.google.com site willoften suggest another search if you didn’tspell one of the keywords correctly.Note 2: Google doesn’t take into consid-

The Googlesearch engineuses a technol-ogy calledPageRank.PageRankrelies on theuniquely demo-cratic nature ofthe web byusing its vastlink structure

as an indicator of an individual page’svalue. In essence, Google interprets alink from page A to page B as a vote,by page A, for page B. But Google looksat more than the sheer volume ofvotes—or links—that a page receives;it also analyzes the page that casts thevote. Votes cast by pages that arethemselves “important” weigh moreheavily and help to make other pages“important.”

However, it is worth understanding twothings about the Google search engine:

Google tends to rank web docu-ments higher than other types of docu-ments. This is simply because web(HTML) documents are more likely to bereferenced by other web documentsthan other types of documents, like pdffiles or Excel spreadsheets. HTMLmakes linking easier and Google usesHTML links to rank pages. An analogymight be that you are more likely toreceive junk mail in your email mailboxthan in your postal mailbox, since it’seasier to advertise a magical dietarysupplement by sending 100,000 emailsthan via the postal services.

The more documents indexed, thebetter the ranking. The official Googlesite (www.google.com) indexes thewhole internet. When indexing a smaller

network, like an intranet, it is more diffi-cult to produce a good ranking, becausethere’s less information available forcalculating the ranks. Like a poll, thebigger the sampling is, the more accu-rate the results.

Google at the DOE JGI

DOE JGI has its own Google searchengine, which indexes documents storedon some local machines (see list below).It can be accessed via http://search.jgi-psf.org/ or via the Search the DOE JGIIntranet box located on the top of pageson the internal website and on the DOEJGI wiki.

More Than Just a Search Engine

The main Google site also providesspecial features to help you find answers.Using www.google.com, you can:

• Evaluate mathematical expres-sions ( 5 + (2 * 2) ).

• Define words (define:species).• Find out about the weather (weather

concord, ca).• Convert units in plain English (16

oz in liter or 2.3 USD per gallon inEUR per liter).

More examples are at http://www.google.com/help/features.html.

Google Advanced Search Operators

The DOE JGI search engine doesn’thave the features we’ve just mentioned(calculator, definition, weather, etc.); how-ever, it does provide advanced searchkeywords that will help DOE JGI usersrefine their searches (these search key-words are also available on www.google.com). Keywords allow you to refine yoursearch to specific types of documents,to specific websites, or to documenttitles only. Here are some useful exam-

cont. on page 11

RENÉ’S TIPS

TIP 3: ADVANCED GOOGLE SEARCHING Understanding Better How Google Works

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 11

eration the capitalization of the key-words (EROOM = eroom = eRoom).

MS Documents Metadata

Metadata (data about data) isattached to each MS document (to seeit, open a MS document and go to File—> Properties). Google is using some ofthese properties for indexing docu-ments. The properties used are:

• Title (can be refined even furtherby using the intitle operator)

• Subject• Author• Keywords

More Tips and Tricks

More Tips and Tricks are availableon Rene Perrier’s twiki page:http://wiki.jgi-psf.org/cgi-bin/twiki/view/Main/RenePerrier

cyanobacterium, Anabaena variabilis,has a unique system of N2 fixation thatinvolves three different nitrogenasegenes—two molybdenum-dependentand one vanadium-dependent. The fixa-tion of atmospheric nitrogen into a formthat plants can readily use is an essen-tial component of the global nitrogencycle, and is of interest in itself. Of par-ticular interest in sequencing A. vari-abilis, however, is the production of H2that occurs as a byproduct of N2 fixa-tion. A better understanding of thegenome of A. variabilis may contribute tothe development of strains capable ofgenerating large amounts of H2 for useas a renewable energy source.

Thiel was open to questions andinput during her seminar, and said morethan once that she was “here to learnas much as to teach.” Following theseminar, Thiel had the opportunity tomeet with many different researchersinvolved in multiple aspects of sequenc-ing and analysis.

Of particular aid to speakers whocome to LANL is comparative analysisof their organism with the genome ofone or more similar or related organ-isms. DOE JGI LANL, in addition toassisting in these analyses, has wel-comed students and other researchersto come and learn how to do their owncomparative analyses.

Many extended collaborations haveformed as a result of the GenomicExplorers Seminar Series. Following thefirst seminar, given by Tom Inzana of theVirginia Polytechnic Institute inDecember 2004, Inzana and JeanChallacombe (JGI LANL) continued towork together on Haemophilus somnus129 Pt, comparing this non-pathogenicbovine commensal to H. ducreyi, a sexu-ally transmitted localized pathogen, andH. influenzae RD, a non-pathogenic lab

strain. They are currently finishing apaper reporting the insights that thesecomparative analyses have providedinto metabolism and host colonization.They also plan to continue their collabo-ration, comparing the non-pathogenicstrain H. somnus 129 Pt to a pathogen-ic strain (H. somnus 2336) that DOE JGILANL just recently finished sequencing.

Another collaboration resulted from aseminar given by Randy Berka in January2005 on the fungus Trichoderma reesei.Berka, who works for Novozymes, Inc.,in Davis, California, and Diego Martinezof DOE JGI LANL expect to publish apaper this year on T. reesei, which is ofparticular interest for its efficient pro-duction of cellulase, an enzyme thatbreaks down cellulose. The fermenta-tion product of cellulose degradationcan be processed to yield fuel-gradeethanol, so a better understanding ofcellulase production may eventuallyhelp lower the cost of cellulose degra-dation and help make bioethanol finan-cially viable as an alternative fuel.

In March, Cindy Nakatsu will cometo LANL from Purdue University inIndiana to give a seminar on Arthrobactersp. FB24, a bacterium with a greatpotential application in bioremediationbecause of its extremely high degree ofchromium tolerance, its resistance tosome other metals and to radiation,and its ability to degrade certain hydro-carbons. Nakatsu’s seminar is guaran-teed to open with the same statementthat Paul Gilna, leader of the GenomicSequencing and Computational BiologyGroup at LANL, makes at the beginningof every seminar: “The DOE JGI is hereto serve the scientific community….”

For more information on theGenomic Explorers Seminar Series, con-tact Jean Challacombe at LANL: jchal-la@lanl.gov.

LANL

René’s Tipscont. from page 10

cont. from page 3

La Fonda on the Plaza, Santa Fe,NM — May 4th and 5th, 2006

In beautiful, historic Santa Fe, NewMexico. Day one of the conferencewill focus on current genome finishingaccomplishments, techniques and pit-falls. Day two will focus on currentand proposed R&D efforts aimed atcutting costs and increasing through-put of the genome finishing process.For more information contact:Chris Detter at (505) 667-1326 orcdetter@lanl.gov

Finishing in the Future

Spring 2006 Vol. 3, Issue 1

12 / THE PRIMER

October 27, 2005To Joni Fazo:Zachary RobertSchmiedt9 lbs/21 inches long

November 11, 2005To Dan Rokhsar (AnnSussman):Abraham SussmanRokhsar7 lbs 1 oz/20 3/4 inches

January 23, 2006To Teresa Green: Abashanti Santee7 lbs 11 oz/22 inches

February 15, 2006To Susannah GreenTringe: Laurel Kathryn Tringe7 lbs 3 oz/19 1/4inches

FUTURE JGI WORKFORCEcomputational tools as CRITICA (CodingRegion Identification Tool InvokingAnalysis) and GLIMMER (Gene Locatorand Interpolated Markov ModelER), theygenerate a master list of genes. Thishelps in visualizing gene positions andresolving trouble points in the initialdraft sequence. The proteins encodedby the found genes are then comparedagainst a microbial sequence databaseusing a set of algorithms contained inthe BLAST (Basic Local AlignmentSearch Tools) suite of programs.

“We use these criteria to resolveoverlaps,” says Loren. “For example, ifyou have a situation where one has aBLAST hit and one doesn’t, you keepthe one with the BLAST hit and throwthe other one away.” In this manner thelist is updated and revised. It is thencompared to the spectrum of annotatedsequences available from other data-bases in an attempt to assign functionsto genes that have been identified with-in the sequence.

“After we go through all the data-base searches and predictions, wehave then a hierarchy of informationthat we give a function call, or what Icall a product description. When you doa BLAST search…you get back your hit:an alignment, a score—the productdescription. So we need to be the mostaccurate with…the product descriptionbecause that’s what everybody seeswhen they do a BLAST job.”

The annotated genome is sent backto the PGF, where the information isuploaded into the Integrated MicrobialGenomes (IMG) data management sys-tem, the DOE JGI portals, and theNational Center for BiotechnologyInformation’s (NCBI’s) GenBank for useby the global research community.

Shewanella is a major microbialplayer in the bioremediation of environ-ments contaminated with toxic metals

and radioactive waste. Its characteriza-tion for the Shewanella Federation, whichis led by Margie Romine at PacificNorthwest National Laboratory (PNNL),illustrates how automation tools canspeed up the annotation process. Underthe auspices of the DOE Office ofBiological and Environmental Research(BER) Microbial Genome SequencingProgram, DOE JGI is contributingsequences from 14 different species ofthe bacterium to this effort.

“Once we annotate as accurately aswe can, we then transfer all of thatinformation directly to the other species,making the initial time investment wellworth it,” says Loren.

The whole process would be quickand easy if only all the microbial organ-isms were structured in a similar way.One of the important aspects of how theydiffer is the GC content, or the propor-tion of G and C bases in the sequence.Genomes with low GC content (below 60percent) can be annotated fairly quicklyusing the available process-automationtools. High GC content genomes stillrequire a significant time investment.

When can you say that a genomehas been fully annotated?

“E. coli is not fully annotated yet,”Loren said. “It is the best annotated,because it has the most experimentaldata, but it is not fully annotated yet.”

“So a lot of the annotation, filling

holes, still depends on future experimen-tal data,” adds Miriam. “That’s one of thebiggest problems right now is that we cangenerate sequence data and get prelim-inary annotation a heck of a lot fasterthan we can provide even high through-put experimental data to verify thatthese things are real, or their function.”

Miriam, Loren, and their colleaguesin the bioinformatics community arestriving to establish a set of rules thatwould allow them to more easilydescribe the features of the genomeand eventually automate the process.According to Miriam, “Hopefully, within ayear or two we will have a list of prod-uct descriptions that would be standardoperating procedure that will apply to allgenomes. The advantage of this is thatall of our genomes will be annotated ina consistent fashion.”

ORNLcont. from page 2

ORNL Annotators Miriam Land and LorenHauser

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 13

BY WENDY SCHACKWITZ

Mourning doves (Zenaida macroura)are a common site at the DOE JGIProduction Genomics Facility. They canbe seen foraging on the ground forgrains and seeds, and perching on thepower lines. They are among the mostabundant birds in North America, andthrive in the environment that we Homosapiens provide. The DOE JGI VertebrateGenomics Program’s Anna Ustaszewskawas lucky enough to have a pair ofmourning doves choose her balcony asa site to raise their babies. I askedAnna to share her experience with us inthis issue’s Birds of DOE JGI article.

There is a beautiful, huge redwoodtree right outside my Walnut Creekapartment. Over the summer, a pair ofmourning doves decided to start a familyon one of its branches. At first, thefemale was just trying out the spot, sit-ting for an hour or two on the branchwhile the male watched over from closeby. Both would then fly away for the restof the day. Doves make a characteristicwhistling noise with their wings whenflying, so it was quite easy to hearthem come and go. After a few days ofthis, they decided to stay and build aflimsy nest of twigs and dry leaves.Soon two eggs appeared. The parentstook turns incubating. After two weeks,the eggs hatched! (At this point, pictureme rolling on the balcony with the cam-era, every chance I got, trying to cap-ture the best view into the nest.)

The young were naked, blind littlethings, unable to hold their heads upfor the first few days. Both parents fedthe young, first with pigeon’s milk, awhite fluid produced in the bird’s cropthat’s similar to the milk of mammals,then solid food. The babies were neverleft unattended in the nest (to the frus-tration of this photographer), except for

brief moments during the parental“switch off.” After two weeks, the youngwere ready to leave the nest. The lastpicture was taken the evening beforethey fledged.

Some facts about mourning doves:Both males and females are about

12 inches in length, with a 28-inchwingspan. They each have a plainbrown, plump body, and a small, pinkishhead with a black spot on the lowercheek. Their long tails taper to a point,and they have black spots on theirupper wings. The male has a small, iri-descent blue patch of feathers on theback of its head and a pinkish breast.

The bird is named for its mournfulcall, “Oowooo-woo-woo,” which is often

mistaken for the hoot of an owl.According to some research, mourn-

ing doves mate for life. They are verysuccessful breeders. Parents may haveseveral broods per year, each consist-ing of two or three chicks.

Their nest is usually a flimsy bowl oftwigs. During nest building, the femalestays at the nest and the male collectssticks. He stands on her back to giveher the nest material. She takes it andweaves it into the nest. Maybe that’swhy the nests are so poorly built!

They are the most widespread andabundant game bird in North America.Despite being hunted throughout mostof their range, they remain among the 10most abundant birds in the United States.

For more information about mourn-ing doves and to listen to their cry, visitCornell’s website: http://www.birds.cor-nell.edu/programs/AllAboutBirds/BirdGuide/Mourning_Dove.html

For more on the evolutionary rela-tionships of birds or any other organ-ism, visit the Tree of Life website:http://tolweb.org/

BIRDS OF DOE JGI

Spring 2006 Vol. 3, Issue 1

14 / THE PRIMER

ing capability at DOE JGILANL and lead the labteam as part of a cooper-ative arrangement betweenthe Walnut Creek andLos Alamos facilities.According to Tom Brettin,Acting Group Leader forthe B-5 GenomicSequencing andComputational BiologyGroup, “Chris is filling acritical position at DOEJGI LANL that has beenvacant since David Bruceaccepted the position ofProject Manager for the DOE JGIMicrobial Program in the fall of 2004.”

One of the goals Chris has set forthe finishing team is to increase the

consistency and efficien-cy of its sequencing pro-cedures. Toward this endthe group is investing innew technology torebuild their infrastruc-ture. For example, a newBiomek FX liquid handlerwas acquired recently tohelp automate sequenc-ing reactions.

“I like technology,”says Chris. “I like mak-ing things bigger, better,and faster.” But in addi-tion to updating equip-

ment to increase efficiency, Chris wantsto improve coordination between thelab team and the computational team.By encouraging the lab team to be

more involved with the computationalanalysis of the raw sequence beforeprotocols are ordered, Chris hopes thatall parties can contribute to decidingwhich protocols are best. This can saveprecious time by eliminating some ofthe redundancy in the process.

“Chris brings significant technicalskills to these high-throughput lab oper-ations,” says Brettin. Chris hopes toshare the knowledge he has amassedat DOE JGI PGF with his new group. Sofar, he has worked to get to know theDOE JGI LANL team members and toevaluate how to achieve his goals.Chris is pleased that his new col-leagues have given him the benefit ofthe doubt and given his ideas and sug-gestions a positive reception.

Although New Mexico is a little coldfor someone who’s spent most of hislife in either Florida or California, Chrisis enjoying the change thus far. LosAlamos is nestled up against the moun-tains, making it a great place for hiking,mountain biking, and skiing. And it’sonly five minutes from work!

He’s even had time to get out andhave some fun. Chris, who’s been ridingmotorcycles since he was ten, quicklyfound fellow riders in B Division withwhom to explore the “Land ofEnchantment.”

Detter Heads Southcont. from page 3

Diatoms areeukaryotic photo-synthetic microor-ganisms found

throughout marine and freshwaterecosystems. Major players in the car-bon cycle, diatoms are responsible forabout 20 percent of global productionof new biomass by photosynthesis.

Phaeodactylum tricornutum is thesecond diatom sequenced. This 30 mil-lion base genome, with the diatom

Thalassiosira pseudonana (alsosequenced by DOE JGI), provides thebasis for comparative genomics studiesand will serve as a foundation for inter-preting the ecological success of theseorganisms.

Forty-plus biologists will gatherMarch 22-24, 2006, at the EmbassySuites, Walnut Creek, Calif., for thePhaeodactylum Jamboree led by theprincipal investigator Chris Bowler ofthe Ecole Normale Superieure and the

DOE JGI annotation team of Alan Kuo,Asaf Salamov, and Igor Grigoriev. OtherJamborees include:

For more information, contact IgorGrigoriev: IVGrigoriev@lbl.gov or (925)296-5860.

SPRING JAMBOREE ROUNDUP

Resources to make the soybean projecta reality.

“Sequencing the soybean genomewill create new opportunities for theadvancement of biodiesel as an econom-ically viable transportation fuel,” saysTuskan. “Having the catalog of all soy-bean genes at our fingertips will facilitatethe discovery of metabolic processes

that will lead to improved oil productionin the seeds and possibly throughout theentire plant. Greater yields of oil per unitarea of land will reduce the cost of pro-duction and move soybean-based bio-diesel closer to your local gas station.”

More information on the LSP can befound at: http://www.jgi.doe.gov/pro-grams/LSP/index.html.

LSP Leadcont. from page 2

Laccariabicolor

April 4-5Nancy,France

Aspergillusniger

April 12-14Vienna,Austria

Xenopustropicalis

April 24-28 Walnut

Creek, Calif.

Spring 2006 Vol. 3, Issue 1

THE PRIMER / 15

“This one was waytoo easy for some-one that has beenwalking around thePGF for over 6 years...” said the identifier of the last Mystery Pic, JGI Facilities manGreg Stanley. If you’ve spotted this one, let gilbert21@llnl.gov know where.

JGI MYSTERY PIC

vest. A collaboration of LANL, severaluniversities, the USDA AgriculturalResearch Service (ARS), and representa-tives from the wine, almond, and citrusindustries, the Plant Pathogen Consortiumwill develop and apply modern technologyto diagnose and combat plant diseasesof concern to the agricultural industry.

According to Project Leader GoutamGupta of the Bioscience Division at LANL,the first step in this process is to createa knowledge base about plant pathogens.The more researchers can learn abouthow the organisms work, the betterequipped they will be to determinepathways for detection and prevention,and to adapt existing technologies tothese problems. Variability, for example,is a major issue. As Gupta explains, someplant pathogens have various strains,and the biggest problem is figuring outwhich strain is affecting the plant.

This is where the DOE Joint GenomeInstitute (DOE JGI) will be especiallyhelpful. Through gene sequencing andannotation, and subsequent analysis,Consortium researchers can develop aknowledge base for these pathogens.Using sequence information and bioin-formatics, along with analysis of path-

ways and gene expression, they canevaluate the specific characteristics thatmake each pathogen unique. Oneapproach may be to distinguish betweenthe strains by looking for specific geneticmarkers and virulence factors. Anothermethod goes one step further and looksat macromolecules such as proteins andcarbohydrates that may be displayed bythe host as early indicators of infection.This kind of investigation can help delin-eate the early steps of plant-pathogeninteractions, which will then lead to ther-apeutic techniques.

The Consortium picked two common,but destructive, organisms to beginresearching: Citrus tristeza virus (CTV)and Xylella fastidiosa (Xf). Citrus tristezavirus is a single-stranded RNA (+) fila-mentous virus that is transmitted byaphids. It is the most destructive virusin citrus trees and manifests in manyways: it can cause the tree to yellowand wilt rapidly (commonly called “quickdecline”) or can result in pitted trunks,stunted growth, and irregular fruit. X.fastidiosa is a gram-negative bacteriumthat is transmitted by insects calledsharpshooters. It infects a variety ofplants, causing numerous diseases,

such as variegated chlorosis in citrustrees, Pierce’s disease in grapevines,and leaf scorch in almond trees. Thesediseases can cause problems such asweakened plants, dead leaves, and dis-figured, tasteless fruit. The Consortiumis already looking at adding a neworganism, citrus greening Liberibacter,to the research plan.

Two X. fastidiosa strains isolated fromdiseased almond trees in the southernSan Joaquin Valley of California wererecently sequenced by DOE JGI.Libraries were compiled at DOE JGILANL, sequencing was done at DOE JGIProduction Genomics Facility, and thefinishing will be completed at DOE JGILANL. These two X. fastidiosa isolateshave different genotypes based on sin-gle nucleotide polymorphisms in the16S DNA. In addition, they exhibit phe-notypic differences through differentcolony-forming properties in vitro.

“The relationships of these variantsto the pathogen biology and diseaseepidemiology are unknown. We expectthat the genomic sequences of thesetwo variant Xf strains will provide insightinto the genetic basis of their biology ormicrobial ecology, as well as their rela-tive roles in the epidemiology of almondleaf scorch disease,” says Gupta.

Identification of key signatures suchas these genotypic variations may be thebest strategy for early detection; waitingfor symptoms could be too late. ThePlant Pathogen Consortium hopes thatby starting with the genetic informationand later studying macromolecules andhost-pathogen relationships, they will beable to develop modern diagnostics andeffective therapeutics for these pathogens.Once the Consortium has successfullydeveloped strategies for these organisms,they will look to expand their investiga-tion to other problem pests.

Plant Pathogen cont. from page 3

Spring 2006 Vol. 3, Issue 1

16 / THE PRIMER

CSO 10993

Tech Transfercont. from page 8

CYBIO COMBINED CHEMISTRYEQUIPMENT (Simon Roberts, SteveWilson, Kecia Duffy-Wei, Simona Necula,and David Robinson)

On December 12 we received thefirst of two combined chemistry instru-ments from CyBio. These machines willreplace the Hydra-Twisters as well as theCavros (some of the PGF’s oldest equip-ment) in the sequencing preparation lab.The CyBio instrument will integrate thetemplate aliquot and the sequencingcocktail dispense into a single automatedrun. In addition, these instruments willembrace a new “web services” technol-ogy from the production informaticsgroup that will improve speed, flexibility,and reliability when communicatingbetween lab instruments and the data-base. Production operators will nolonger hand-scan plate barcodes into

the database, availing more time to multi-task. The CyBio instruments will eventuallyutilize vendor applied barcodes, elimi-nating the need for operators to hand-apply printed barcodes. The first CyBioCombined Chemistry instrument wasmoved into the production lab 140 and

began daily processing of productionplates on a limited scale on February22. Production scale-up of the firstinstrument will continue through March2006. The second CyBio CombinedChemistry instrument is scheduled tocome online in April 2006.

The CyBio CyBi-Well Vario and CyBi-Drop height-adjustable table was pur-chased from Lista Toolboxes in MenloPark, CA, with a 21-inch MIN to 31-inchMAX height adjustment range. This wasspecified to give an ergonomic workingheight for all operators, both short andtall, based on an Ergonomic RiskReduction Analysis called STEER. Thestandard fixed lab bench section wasremoved to install the system andtable. With casters on the table, it alsohas the advantage of enabling easyaccess for maintenance if required.

CONTACT THE PRIMER / David Gilbert, Editor / gilbert21@llnl.gov / (925) 296-5643

PGF senior research tech David Robinsonhas a new addition to his workspace, thetime-saving Cybio instrument.

New SeniorRecruiter BillCannan hasjoined the DOEJGI HumanResourcesteam. In thisrole, he will pro-vide comprehensive recruitment,staffing, and outsourcing support,including representing the JGI in vari-ous recruitment efforts. Bill’s recruit-ing experience includes over eightyears with California State AutomobileAssociation, West Valley Engineering,and Kelly Services Onsite @ Lifescan.He brings strong knowledge ofCalifornia employment labor laws,demonstrated success in high-volumerecruiting, and over thirteen years ofclient service experience.

CSP PROJECTS IN PROGRESS

The tropical grain Sorghum bicolor, proposed by an inter-national consortium led by the University of Georgia andRutgers, will complement the knowledge gleaned from rice,the only other monocot grain sequenced to date. The U.S.provides 70 to 80 percent of world sorghum exports. Theworldwide annual economic value of the crop exceeds$69 billion. With a relatively compact genome of approxi-mately 736 million bases, Sorghum will serve as a valuablereference for analyzing the fourfold larger genome of maize,the leading U.S. fuel ethanol crop. About 12 percent ofthe U.S. grain sorghum crop goes into ethanol production.Sorghum is an even closer relative of sugarcane, arguablythe most important biofuels crop worldwide.

The 2007 DOEJGI Community

SequencingProgram call forLetters of Intent

generated:

234 letters of intent which break down into:

• 25 large genomes/8 given the go-ahead for fullproposal submission

• 118 microbes/88

• 91 small eukaryotic genome projects/65

147 full proposals were received on March 10and will be reviewed in early May.