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Biotechniques. 2001 Jun;30(6):1250-4.

Cloning full-length, cap-trapper-selected cDNAs by using the single-strand linker ligation method.

Author information

1
RIKEN Tsukuba Institute, Ibaraki, Japan.

Abstract

We have developed the single-strand linker ligation method (SSLLM), which uses DNA ligase to add a dsDNA linker to single-stranded (ss) full-length cDNA. The linkers have random 6-bp (dN6 or dGN5) 3' overhangs that can ligate to any cDNA sequence, thereby facilitating the production of cDNA libraries with titers exceeding 1 x 10(6) independent clones. We confirmed that the 5' ends of cDNA inserts cloned by using SSLLM are full-length and include the 5' untranslated regions. The great advantage of our method is that the elimination of the GC tail simplifies the sequencing and protein translation of the full-length clones. Further, our method tags ss cDNAs more efficiently than does the traditional RNA ligase reaction.

PMID:
11414214
DOI:
10.2144/01306st01
[Indexed for MEDLINE]
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