DOE Joint Genome Institute

  • COVID-19
  • About Us
  • Contact Us
  • Our Science
    • DOE Mission Areas
    • Bioenergy Research Centers
    • Science Programs
    • Science Highlights
    • Scientists
    (PXFuel)
    Designer DNA: JGI Helps Users Blaze New Biosynthetic Pathways
    In a special issue of the journal Synthetic Biology, JGI scientific users share how they’ve worked with the JGI DNA Synthesis Science Program and what they’ve discovered through their collaborations.

    More

    A genetic element that generates targeted mutations, called diversity-generating retroelements (DGRs), are found in viruses, as well as bacteria and archaea. Most DGRs found in viruses appear to be in their tail fibers. These tail fibers – signified in the cartoon by the blue virus’ downward pointing ‘arms’— allow the virus to attach to one cell type (red), but not the other (purple). DGRs mutate these ‘arms,’ giving the virus opportunities to switch to different prey, like the purple cell. (Courtesy of Blair Paul)
    A Natural Mechanism Can Turbocharge Viral Evolution
    A team has discovered that diversity generating retroelements (DGRs) are not only widespread, but also surprisingly active. In viruses, DGRs appear to generate diversity quickly, allowing these viruses to target new microbial prey.

    More

    Algae growing in a bioreactor. (Dennis Schroeder, NREL)
    Refining the Process of Identifying Algae Biotechnology Candidates
    Researchers combined expertise at the National Labs to screen, characterize, sequence and then analyze the genomes and multi-omics datasets for algae that can be used for large-scale production of biofuels and bioproducts.

    More

  • Our Projects
    • Search JGI Projects
    • DOE Metrics/Statistics
    • Approved User Proposals
    • Legacy Projects
    This data image shows the monthly average sea surface temperature for May 2015. Between 2013 and 2016, a large mass of unusually warm ocean water--nicknamed the blob--dominated the North Pacific, indicated here by red, pink, and yellow colors signifying temperatures as much as three degrees Celsius (five degrees Fahrenheit) higher than average. Data are from the NASA Multi-scale Ultra-high Resolution Sea Surface Temperature (MUR SST) Analysis product. (Courtesy NASA Physical Oceanography Distributed Active Archive Center)
    When “The Blob” Made It Hotter Under the Water
    Researchers tracked the impact of a large-scale heatwave event in the ocean known as “The Blob” as part of an approved proposal through the Community Science Program.

    More

    A plantation of poplar trees. (David Gilbert)
    Genome Insider podcast: THE Bioenergy Tree
    The US Department of Energy’s favorite tree is poplar. In this episode, hear from ORNL scientists who have uncovered remarkable genetic secrets that bring us closer to making poplar an economical and sustainable source of energy and materials.

    More

    Ian Rambo, graduate student at UT-Austin, was a DOE Graduate Student Research Fellow at the JGI
    Virus-Microbe Interactions of Mud Island Mangroves
    Through the DOE Office of Science Graduate Student Research (SCGSR) program, Ian Rambo worked on part of his dissertation at the JGI. The chapter focuses on how viruses influence carbon cycling in coastal mangroves.

    More

  • Data & Tools
    • IMG
    • Data Portal
    • MycoCosm
    • PhycoCosm
    • Phytozome
    • GOLD
    HPCwire Editor's Choice Award (logo crop) for Best Use of HPC in the Life Sciences
    JGI Part of Berkeley Lab Team Awarded Best Use of HPC in Life Sciences
    The HPCwire Editors Choice Award for Best Use of HPC in Life Sciences went to the Berkeley Lab team comprised of JGI and ExaBiome Project team, supported by the DOE Exascale Computing Project for MetaHipMer, an end-to-end genome assembler that supports “an unprecedented assembly of environmental microbiomes.”

    More

    With a common set of "baseline metadata," JGI users can more easily access public data sets. (Steve Wilson)
    A User-Centered Approach to Accessing JGI Data
    Reflecting a structural shift in data access, the JGI Data Portal offers a way for users to more easily access public data sets through a common set of metadata.

    More

    Phytozome portal collage
    A More Intuitive Phytozome Interface
    Phytozome v13 now hosts upwards of 250 plant genomes and provides users with the genome browsers, gene pages, search, BLAST and BioMart data warehouse interfaces they have come to rely on, with a more intuitive interface.

    More

  • User Programs
    • Calls for Proposals
    • Special Initiatives & Programs
    • Product Offerings
    • User Support
    • Policies
    • Submit a Proposal
    screencap from Amundson and Wilkins subsurface microbiome video
    Digging into Microbial Ecosystems Deep Underground
    JGI users and microbiome researchers at Colorado State University have many questions about the microbial communities deep underground, including the role viral infection may play in other natural ecosystems.

    Read more

    Yeast strains engineered for the biochemical conversion of glucose to value-added products are limited in chemical output due to growth and viability constraints. Cell extracts provide an alternative format for chemical synthesis in the absence of cell growth by isolating the soluble components of lysed cells. By separating the production of enzymes (during growth) and the biochemical production process (in cell-free reactions), this framework enables biosynthesis of diverse chemical products at volumetric productivities greater than the source strains. (Blake Rasor)
    Boosting Small Molecule Production in Super “Soup”
    Researchers supported through the Emerging Technologies Opportunity Program describe a two-pronged approach that starts with engineered yeast cells but then moves out of the cell structure into a cell-free system.

    More

    These bright green spots are fluorescently labelled bacteria from soil collected from the surface of plant roots. For reference, the scale bar at bottom right is 10 micrometers long. (Rhona Stuart)
    A Powerful Technique to Study Microbes, Now Easier
    In JGI's Genome Insider podcast: LLNL biologist Jennifer Pett-Ridge collaborated with JGI scientists through the Emerging Technologies Opportunity Program to semi-automate experiments that measure microbial activity in soil.

    More

  • News & Publications
    • News
    • Blog
    • Podcasts
    • Webinars
    • Publications
    • Newsletter
    • Logos and Templates
    • Photos
    In their approved proposal, Frederick Colwell of Oregon State University and colleagues are interested in the microbial communities that live on Alaska’s glacially dominated Copper River Delta. They’re looking at how the microbes in these high latitude wetlands, such as the Copper River Delta wetland pond shown here, cycle carbon. (Courtesy of Rick Colwell)
    Monitoring Inter-Organism Interactions Within Ecosystems
    Many of the proposals approved through JGI's annual Community Science Program call focus on harnessing genomics to developing sustainable resources for biofuels and bioproducts.

    More

    Coloring the water, the algae Phaeocystis blooms off the side of the sampling vessel, Polarstern, in the temperate region of the North Atlantic. (Katrin Schmidt)
    Climate Change Threatens Base of Polar Oceans’ Bountiful Food Webs
    As warm-adapted microbes edge polewards, they’d oust resident tiny algae. It's a trend that threatens to destabilize the delicate marine food web and change the oceans as we know them.

    More

    Integrating JGI Capabilities for Exploring Earth’s Secondary Metabolome
    Natural Prodcast podcast: Nigel Mouncey
    JGI Director Nigel Mouncey has a vision to build out an integrative genomics approach to looking at the interactions of organisms and environments. He also sees secondary metabolism analysis and research as a driver for novel technologies that can serve all JGI users.

    More

Our Science
Home › Our Science › Science Programs › Plant Program › Brachypodium Resources › Brachypodium T-DNA Collection

Brachypodium T-DNA Collection

Formerly the WRRC Brachypodium distachyon T-DNA collection.

For information about sequenced chemical and radiation mutants and other resources please visit the Brachypodium Resources site

Current collection statistics

Last Updated: 5-28-2019

  • Lines in T-DNA collection: 23,649
  • Unique insertions sites identified: 25,977

Project description

The small grass species Brachypodium distachyon (Brachypodium) combines the desirable attributes of a model organism with many of the traits of interest for the development and improvement of grasses (such as wheat, barley, switchgrass, and Miscanthus giganteus) that are of worldwide importance as sources of food, feed and fuel. In recognition of its utility, the DOE called for development of Brachypodium as a model plant for use in the domestication of energy crops in the 2005 report “Breaking the Biological Barriers to Cellulosic Ethanol: A Joint Research Agenda.” Since then, a comprehensive infrastructure of genomic resources has been assembled for Brachypodium.

The goal of this project (funded by the DOE Feedstock Genomics Program through interagency agreement # 60-5325-7-573) is to add to the growing collection of genomic resources available for Brachypodium by creating a large collection of T-DNA lines. These lines are indexed through flanking sequence tags (FSTs) that facilitate mapping of the T-DNA insertions within the Brachypodium genome. The collection can therefore serve to identify mutations in genes predicted to affect biomass quality and agronomic characteristics of cereal and energy crops. This paper describes the creation and sequencing of the first 8,491 T-DNA lines. This paper describes the generation and sequencing of the rest of the collection using TDNA-Seq.

  • T-DNA insertions Bd21 V3 assembly 3-18-2016 (updated to the V3.o genome assembly)
  • Search for insertions in specific genes using jbrowse in the Bd21 genome assembly or in the Bd21-3 genome assembly. The mutants are in line Bd21-3 but the original reference genome was made from line Bd21.

Collection details

This collection comprises plants produced using a variety of constructs designed for different purposes. All lines are T-DNA insertional mutants, and therefore have the potential to create gene knockouts, and this is the exclusive purpose of lines produced using the pOL001, pJJH, pJJ2LB, pJJB, and pJJB2LB vectors. In addition, the T-DNAs of the pJJ2LBP and pJJ2LBP2 vectors contain “gene trap” sequences. In these vectors, a promotor-less GUS gene is placed adjacent to the left border, and a promotor-less GFP gene resides adjacent to the right border. If the T-DNA integrates downstream of a promoter, expression of one of the reporter genes could be used to infer the expression pattern of the disrupted gene. These constructs also contains multiple splice acceptor sites adjacent to the reporter genes to allow efficient splicing should the T-DNA fall into an intron. These lines have the potential to provide clues about the role of the disrupted gene, to provide a tool to understand the function of the disrupted gene, and to identify promoters with useful expression patterns.

The pJJ2LBA and pJJ2LBA2 vectors contain transcriptional enhancers within the T-DNA sequence. These “activation tagging” constructs are designed to increase the transcription of nearby genes. Importantly, the transcriptional enhancers are designed to give overexpression with the same expression pattern, rather than constitutive expression, of affected genes. Activation tagging is particularly well suited to assign function to genes with redundant functions where knockouts in an individual family member do not produce a phenotype.

In generating this collection, we have done an extensive evaluation of vectors built with different promoters, reporter genes, and selectable markers in order to optimize transformation efficiency. We have observed considerable variation in efficiency and plant survival and fertility depending on the construct used. Our observations include the following: Transformations employing hygromycin selection yielded consistently higher efficiency and survival over those using BASTA. The promoter driving the selectable marker greatly affects transformation efficiency (maize ubiquitin > CaMV 35S with a 5′ intron> CaMV 35S without a 5′ intron >> rice tubulin). Considering time and ease of evaluation, screening transformed tissue for GUS staining was more efficient than checking for GFP or RFP fluorescence. T-DNA vectors containing two left border sequences produce transformants that yield a higher rate of successfully recovering sequence flanking the T-DNA insertion sites. Ac/Ds and En/Spm transposons function in Brachypodium, but are lethal, possibly because they are too active.

Vector sequences:

  • pJJ2LB
  • pJJ2LBA2_and_pJJ2LBA
  • pJJ2LBP2_and_pJJ2LBP
  • pOL001

Citations

For T-DNA mutants with flanking sequences beginning with the prefix “JJ” please cite:

Bragg, J.N., Wu, J., Gordon, S.P., Guttman, M.A., Thilmony, R.L., Lazo, G.R., Gu, Y.Q., Vogel, J.P. 2012 Generation and Characterization of the Western Regional Research Center Brachypodium T-DNA Insertional Mutant Collection PLoS ONE 7 (9) art. no. e41916

For T-DNA mutants with flanking sequences beginning with the prefix “IL” please cite:

Hsia, M.M., O’Malley, R., Cartwright, A., Nieu, R., Gordon, S.P., Sandra Kelly, K., Williams, T.G., Wood, D.F., Zhao, Y., Bragg, J., Jordan, M., Markus Pauly, M., Joseph R. Ecker, J.R., Yong Gu, Y., Vogel, J.P., 2017 Sequencing and functional validation of the JGI Brachypodium distachyon T-DNA collection.  Plant Journal 91(3): p. 361-370.

Ordering Brachypodium T-DNA lines from the JGI Brachypodium collection

Update 5-28-2019: Seed distribution is being transitioned to the USDA National Plant Germplasm System (NPGS). During this transition, we will be unable to fill new orders for seeds. Existing orders will be filled. Seeds are being shipped to NPGS now, but it may be some time before they are available for ordering. We will update this section once seeds start becoming available. You can also search NGPS at  https://www.ars-grin.gov/npgs/ to check availability. Questions can be directed to: BrachypodiumTDNA@gmail.com

Note that we are still distributing seeds for sequenced chemical and radiation mutants. Please visit the Brachypodium Resources site for more information about these mutants.

Download this file for more information about how to genotype T-DNA lines.

 

Diagram of constructs used:
tdna-constructs-diagram

Information about constructs needed for completing a request for notification 

construct pOL001

Phenotypic Designation Name: constitutive GUS expression

Identifying Line(s): ENTER THE LINES REQUESTED HERE

Construct(s): pOL001

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – Visual marker

MG – Hygromycin resistant

Genotype(s)

Selectable Marker

Screenable Marker

Construct components

Promoter: 35S promoter from Cauliflower mosaic caulimovirus – CaMV 35S constitutive promoter to drive expression of hygromycin phosphotransferase selectable marker

Gene: Hygromycin selectable marker from Escherichia coli – hygromycin phosphotransferase gene to confer hygromycin resistance as a selectable marker

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of hygromycin phosphotransferase gene

 

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive expression of GUS reporter gene

Gene: glucuronidase (GUS) reporter gene from Escherichia coli – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GUS gene

constructs pJJH and pJJ2LB

Phenotypic Designation Name: constitutive GUS expression

Identifying Line(s): ENTER THE LINES REQUESTED HERE

Construct(s): pJJH or pJJ2LB

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – Hygromycin resistant

 

Genotype(s)

Selectable marker

Screenable Marker

 

Construct components

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive

expression of hygromycin phosphotransferase selectable marker

Gene: Hygromycin selectable marker from Escherichia coli – hygromycin phosphotransferase gene to confer hygromycin resistance as a selectable marker

 

Promoter: 35S promoter from Cauliflower mosaic caulimovirus – CaMV 35S constitutive promoter to drive expression of GUS reporter gene

Gene: glucuronidase (GUS) reporter gene from Escherichia coli – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GUS gene

constructs pJJB and pJJB2LB

Phenotypic Designation Name: constitutive GUS expression

Identifying Line(s): ENTER THE LINES REQUESTED HERE

 

Construct(s): pJJB or pJJB2LB

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – BASTA resistant

 

Genotype(s)

Selectable marker

Screenable Marker

 

Construct components

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive

expression of phosphinothricin acetyltransferase selectable marker

Gene: BASTA selectable marker from Streptomyces hygrosopicus- phosphinothricin acetyltransferase gene to confer BASTA resistance as a selectable marker

 

Promoter: 35S promoter from Cauliflower mosaic caulimovirus – CaMV 35S constitutive promoter to drive expression of GUS reporter gene

Gene: glucuronidase (GUS) reporter gene from Escherichia coli – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GUS gene

construct pJJ2LBP2

Phenotypic Designation Name: promoter trapping construct

Identifying Line(s): ENTER THE LINES REQUESTED HERE

Construct(s): pJJ2LBP2

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – Hygromycin resistant

Genotype(s)

Selectable Marker

promoterless reporter GUS

promoterless reporter GFP

 

Construct components

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive

expression of hygromycin phosphotransferase selectable marker

Gene: Hygromycin selectable marker from Escherichia coli – hygromycin phosphotransferase gene to confer

hygromycin resistance as a selectable marker

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of hygromycin phosphotransferase gene

 

Gene: glucuronidase (GUS) reporter gene from Escherichia coli – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GUS gene

 

Gene: green fluorescent protein (GFP) reporter gene from Aequorea Victoria – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GFP gene

construct pJJ2LBA2

Phenotypic Designation Name: activation tagging construct

Identifying Line(s): ENTER THE LINES REQUESTED HERE

Construct(s): pJJ2LBA2

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – Hygromycin resistant

Genotype(s)

Selectable Marker

Enhancer element

 

Construct components

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive

expression of hygromycin phosphotransferase selectable marker

Gene: Hygromycin selectable marker from Escherichia coli – hygromycin phosphotransferase gene to confer

hygromycin resistance as a selectable marker

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of hygromycin phosphotransferase gene

 

Enhancer: 4x 35S enhancer element from Cauliflower mosaic caulimovirus – CaMV 35S

construct pJJ2LBP

Phenotypic Designation Name: promoter trapping construct

Identifying Line(s): ENTER THE LINES REQUESTED HERE

Construct(s): pJJ2LBP2

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – Hygromycin resistant

Genotype(s)

Selectable Marker

promoterless reporter GUS

promoterless reporter GFP

 

Construct components

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive

expression of hygromycin phosphotransferase selectable marker

Gene: Hygromycin selectable marker from Escherichia coli – hygromycin phosphotransferase gene to confer

hygromycin resistance as a selectable marker

 

Gene: glucuronidase (GUS) reporter gene from Escherichia coli – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GUS gene

 

Gene: green fluorescent protein (GFP) reporter gene from Aequorea Victoria – reporter gene

3`UTR: nos3′ UTR from Agrobacterium tumefaciens – terminator at end of GFP gene

construct pJJ2LBA

Phenotypic Designation Name: activation tagging construct

Identifying Line(s): ENTER THE LINES REQUESTED HERE

Construct(s): pJJ2LBA2

Mode of Transformation: Agrobacterium tumefaciens, disarmed

 

Phenotype(s)

MG – Hygromycin resistant

Genotype(s)

Selectable Marker

Enhancer element

 

Construct components

Promoter: maize ubiquitin promoter from Zea mays – constitutive promoter to drive

expression of hygromycin phosphotransferase selectable marker

Gene: Hygromycin selectable marker from Escherichia coli – hygromycin phosphotransferase gene to confer

hygromycin resistance as a selectable marker

 

Enhancer: 4x 35S enhancer element from Cauliflower mosaic caulimovirus – CaMV 35S

 

 

For more information contact : brachypodiumTDNA@gmail.com

 

 

 

  • Careers
  • Contact Us
  • Events
  • User Meeting
  • MGM Workshops
  • Internal
  • Disclaimer
  • Credits
  • Policies
  • Emergency Info
  • Accessibility / Section 508 Statement
  • Flickr
  • LinkedIn
  • RSS
  • Twitter
  • YouTube
Lawrence Berkeley National Lab Biosciences Area
A project of the US Department of Energy, Office of Science

JGI is a DOE Office of Science User Facility managed by Lawrence Berkeley National Laboratory

© 1997-2022 The Regents of the University of California