characteristics.text: SAMN00154406
characteristics.characteristic: External Id
characteristics.text: LIBEST_020989
characteristics.characteristic: External Id
characteristics.text: Colorado State University
characteristics.characteristic: INSDC center name
characteristics.text: 2007-05-25T00:00:00Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-10-11T17:45:06.010Z
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: Plant tissue was shipped in dry ice to Agencourt Bioscience Corporation (Beverly MA). Total RNA was extracted and optimized first strand cDNA synthesis was performed using a primer adapted with a rare enzyme cut site. cDNA fragments were size-selected by agarose gel electrophoresis, and directionally cloned into a pAGEN-1 vector. A positive control containing the TetR gene was used during construction of the primary library to ensure library quality. Single stranded DNA was made from a portion of the primary library by phagemid production, and reactions were treated with DNase I to ensure the removal of double stranded DNA. A second portion of the primary library was linearized and transcribed into anti-sense RNA with biotinylated dNTPs. Oligo dT and primer extension were used to pre-block the poly-A region prior to hybridization. The anti-sense RNA and single stranded circular DNA were hybridized, and abundant clones were removed using streptavidin. To reduce the amount of empty vectors, a Not1 oligo and Taq polymerase were used to synthesize double stranded DNA from the single stranded normalized library prior to final transfection. The normalized library was plated and 4969 clones were randomly selected for sequencing. Automated plasmid purification was achieved using the SPRI (SprintPrep) technique, which harvests plasmid DNA directly from lysed bacterial cultures, trapping both plasmid and genomic DNA to functionalized bead particles and selectively eluting only the plasmid (Beckman Biomek FX robots and CCS Packard DNATraks). DNA templates were sequenced in 384-well format using BigDye Version 3.1 reactions on ABI3730 instruments at Agencourt Biosciences. Thermal cycling was performed using 384-well Thermal cyclers (ABI, MJ Research). Sequencing Reactions were purified using Agencourt's CleanSeq dye-terminator removal kit.
characteristics.characteristic: description
characteristics.text: Entire Plant (rosette and roots)
characteristics.characteristic: organ
characteristics.text: Centaurea maculosa
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_215693
characteristics.characteristic: organism
characteristics.text: Centaurea normalized cDNA library
characteristics.characteristic: title
characteristics.text: pAGEN (Agencourt Biosciences)
characteristics.characteristic: v_type
characteristics.text: plasmid
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/SO_0000155
characteristics.characteristic: vector