Linked Life Data

15786810

Source:http://linkedlifedata.com/resource/pubmed/id/15786810

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
2005-3-24
pubmed:abstractText
In this study, we developed a method that allows cDNA library construction from a small amount of RNA without causing serious size bias in the resulting cDNA population. For this purpose, we adopted two-round cRNA amplification by T7 and SP6 RNA polymerases. The first-round cDNAs, flanked by the promoter sequences of T7 and SP6 RNA polymerases, were synthesized from 1 microg total RNA and then subjected to two rounds of cRNA amplification. Comparison of the sizes of the first-round and the second-round cRNAs indicated that the size-bias effect of the second-round cRNA synthesis was not serious. The resultant double-stranded cDNAs were cloned into a plasmid by in vitro lambda phage recombination with an efficiency of 1.2 x 10(11) colony-forming unit/microgram of starting total RNA. Characterization of the resultant cDNA library in terms of the insert size, clone redundancy, and integrity of 3' ends of cDNAs indicated that the amplified library was comparable to a library constructed by a conventional method, although large cDNAs tend to be slightly truncated in the amplified library. This method enables the construction of a library from a small amount of RNA, and calculations suggest that the strategy would be efficient enough to use even a single cell as starting material.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0736-6205
pubmed:author
pubmed:issnType
Print
pubmed:volume
38
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
451-8
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2005
pubmed:articleTitle
CDNA library construction from a small amount of RNA: adaptor-ligation approach for two-round cRNA amplification using T7 and SP6 RNA polymerases.
pubmed:affiliation
Department of Human Gene Research, Kazusa DNA Research Institute, Kisarazu, Chiba, Japan. ohreiko@kazusa.or.jp
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't, Evaluation Studies