Construction of Long-Transcript Enriched cDNA Libraries from Submicrogram Amounts of Total RNAs by a Universal PCR Amplification Method
| Autor: | Meng K. Lim, Minoru S.H. Ko, Yulan Piao, Naomi T. Ko |
|---|---|
| Rok vydání: | 2001 |
| Předmět: |
DNA
Complementary cDNA library Molecular Sequence Data Gene Amplification RNA Biology Polymerase Chain Reaction Molecular biology Insert (molecular biology) law.invention Mice Inbred C57BL Mice Rapid amplification of cDNA ends Start codon law Methods Genetics Animals Genomic library Primer (molecular biology) Genetics (clinical) Polymerase chain reaction Gene Library |
| Zdroj: | Genome Research. 11:1553-1558 |
| ISSN: | 1549-5469 1088-9051 |
| DOI: | 10.1101/gr.185501 |
| Popis: | Here we report a novel design of linker primer that allows one to differentially amplify long tracts (average 3.0 kb with size ranges of 1–7 kb) or short DNAs (average 1.5 kb with size ranges of 0.5–3 kb) from a complex mixture. The method allows one to generate cDNA libraries enriched for long transcripts without size selection of insert DNAs. One representative library from newborn kidney includes 70% of clones bearing ATG start codons. A comparable library has been generated from 20 mouse blastocysts, containing only ∼40 ng of total RNA. This universal PCR amplification scheme can provide a route to isolate very large cDNAs, even if they are expressed at very low levels.[The sequence data described in this paper have been submitted to the GenBank data library under accession numbersBG060207–BG062928.] |
| Databáze: | OpenAIRE |
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