characteristics.text: SAMN00155039
characteristics.characteristic: External Id
characteristics.text: LIBEST_000428
characteristics.characteristic: External Id
characteristics.text: Michigan State University
characteristics.characteristic: INSDC center name
characteristics.text: 1995-08-16T00:00:00Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-10-11T17:35:11.057Z
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: Using 5 ug of polyadenylated mRNA from 3 day-old Arabidopsis thaliana (Columbia) seedling hypocotyls as template and oligo d(t) as primer, first strand synthesis was catalyzed by Moloney murine leukemia virus reverse transcriptase (Pharmacia). Second-strand cDNA was made using the procedure of Gubler and Hoffman (1983) except that DNA ligase was omitted. After the second strand reaction, the ends of the cDNA were made blunt with Klenow fragment and EcoRI/NotI adapters (Pharmacia) were ligated to each end. The cDNA was purified from unligated adapters by spun-column chromatography using sephacryl s-300 and size-fractionated on a 1% low melting point mini-gel. Size selected cDNAs (3 - 6 kb) were removed from the gel using agarase (New England Biolabs), phenol:choloroform extracted and precipitated using 0.3 M NaOAc (pH 7)/ethanol. A portion of each cDNA size-fraction (0.1 ug) was co-precipitated with 1 ug of lambdaZapII (StratageneEcoRI digested, dephosphorylated arms and then ligated in a volume of 4 ul overnight. Each ligation mix was packaged in vitro using Gigapack II gold packaging extract (Stratagene). We have determined that although first strand cDNA synthesis was initiated using dT, almost all of the cDNAs begin 8-10 bp from the poly-A tail. The reason for the loss of the poly-A tail is most likely due to lower than anticipated nucleotide levels during the Klenow repair of ragged ends before the addition of linkers (3'-5' exo instead of 5'-3' pol). When this library is used please reference the ABRC and: Kieber, J. et al. (1993) Cell 72:427-441.
characteristics.characteristic: description
characteristics.text: 3 day-old
characteristics.characteristic: dev-stage
characteristics.text: Arabidopsis thaliana
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_3702
characteristics.characteristic: organism
characteristics.text: EcoRI
characteristics.characteristic: re_1
characteristics.text: EcoRI
characteristics.characteristic: re_2
characteristics.text: Columbia
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0005147
characteristics.characteristic: strain
characteristics.text: seedling hypocotyl
characteristics.characteristic: tissue-type
characteristics.text: CD4-16
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0005180
characteristics.characteristic: title
characteristics.text: pBluescript SK-
characteristics.characteristic: vector