characteristics.text: LIBEST_022337
characteristics.characteristic: External Id
characteristics.text: SAMN00164572
characteristics.characteristic: External Id
characteristics.text: University of California
characteristics.characteristic: INSDC center name
characteristics.text: 2007-12-11T00:00:00Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-10-11T17:45:41.717Z
characteristics.characteristic: INSDC last update
characteristics.text: live
characteristics.characteristic: INSDC status
characteristics.text: Generic
characteristics.characteristic: NCBI submission model
characteristics.text: Generic.1.0
characteristics.characteristic: NCBI submission package
characteristics.text: PI 418979
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CHEBI_90524
characteristics.characteristic: cultivar
characteristics.text: Seeds of PI 418979 were from a single plant with a long history of single seed descent (Ehlers) from China. The types of tissue were: 1) germinated seed hypocotyl and primary root, 2) germinated seed epicotyl and foliage leaves, 3 & 4) young trifoliate leaves and apical meristem (one well watered, one drought stressed). For germinated seed samples (1 & 2), seeds were surface sterilized 5 minutes in 1% sodium hypochlorite with 0.01% Tween 20, rinsed with water, germinated on sterile filter paper with water containing 100 units/ml nystatin and 50 microgram/ml cefotaxime, incubated 4-5 days in the dark at room temperature (Fenton). For samples 3 & 4, seeds were planted in 3 gallon pots and kept watered in a glasshouse until plants were ~25 cm in height (~3 weeks)(Fenton). For drought stress (sample 4), water was withheld until visible signs of drought stress occurred (reduced growth relative to watered plants, wilted leaves, less green color), generally ~3 weeks. Total RNA was purified using a TRIzol method, except Qiagen RNeasy for hypocotyls & root (Fenton). RNA was pooled to establish one complex RNA sample with a total of 500 microgram, as follows: 167 microgram of hypocotyl & primary root (sample 1), 167 microgram of epicotyl and foliage leaves (sample 2) and 87 microgram each of stressed and unstressed young trifoliate leaves and apical meristem (samples 3 & 4)(Fenton). PolyA RNA was purified using PolyATrack mRNA Isolation System IV (Promega)(Fenton). 2.0 microgram of poly(A) mRNA was used for cDNA synthesis. A primary cDNA library with a titer of 15 million cfu (Close) was produced using a SuperScript Plasmid System with Gateway Technology for cDNA Synthesis and Cloning (Invitrogen)(Fenton). Plating and sequencing was performed at the US Department of Energy Joint Genome Institute Sequence Production Facility, Walnut Creek California (Richardson, Lucas, Rokhsar, Wang, Lindquist). The library was plated onto agarose plates containing ampicillin (254 mm plates from Teknova, Hollister, CA) at a density of approximately 1000 colonies per plate. Plates were grown at 37 degrees Celsius for 18 hours, then individual colonies were picked and each used to inoculate a well containing LB media with ampicillin in a 384 well plate (Nunc, Rochester, NY). 384 well plates were grown at 37 degrees Celsius for 18 hours. Plasmid DNA for sequencing was produced by rolling circle amplification (Templiphi, GE Healthcare, Piscataway, NJ). Subclone inserts were sequenced from both ends using primers complementary to the flanking pCMV-SPORT6 vector sequence and Big Dye terminator chemistry and then run on ABI 3730 instruments (ABI, Foster City, CA). Sequencing protocols can be found in Detter et al (2002, Genomics 80:691-698) and at http://www.jgi.doe.gov/sequencing/protocols/index.html. Trace files were downloaded (Close) and then processed (Wanamaker) using the HarvEST pipeline (http://harvest.ucr.edu) to remove vector and cloning oligo sequences and various contaminants, and to trim to a high quality region. Sequences that retained a phred 17 region of at least 100 bases were deposited to GenBank with annotations (Close).
characteristics.characteristic: description
characteristics.text: Seedling
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PO_0007131
characteristics.characteristic: dev-stage
characteristics.text: E. coli DH10B
characteristics.characteristic: lab-host
characteristics.text: Vigna unguiculata
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_3917
characteristics.characteristic: organism
characteristics.text: SalI
characteristics.characteristic: re_1
characteristics.text: NotI
characteristics.characteristic: re_2
characteristics.text: mixed
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0001271
characteristics.characteristic: tissue-type
characteristics.text: Cowpea PI 418979 Mixed Tissue and Conditions cDNA Library UCRVU10-1
characteristics.characteristic: title
characteristics.text: Plasmid
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/SO_0000155
characteristics.characteristic: v_type
characteristics.text: pCMV-SPORT6
characteristics.characteristic: vector