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Anal Biochem. 2013 Oct 15;441(2):101-3. doi: 10.1016/j.ab.2013.06.013. Epub 2013 Jul 4.

A tailing genome walking method suitable for genomes with high local GC content.

Author information

1
State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Public Health of Agricultural Ministry, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China.

Abstract

The tailing genome walking strategies are simple and efficient. However, they sometimes can be restricted due to the low stringency of homo-oligomeric primers. Here we modified their conventional tailing step by adding polythymidine and polyguanine to the target single-stranded DNA (ssDNA). The tailed ssDNA was then amplified exponentially with a specific primer in the known region and a primer comprising 5' polycytosine and 3' polyadenosine. The successful application of this novel method for identifying integration sites mediated by φC31 integrase in goat genome indicates that the method is more suitable for genomes with high complexity and local GC content.

KEYWORDS:

GC content; Genome walking; Goat genome; Integration sites; Tailing; φC31 integrase

PMID:
23831478
DOI:
10.1016/j.ab.2013.06.013
[Indexed for MEDLINE]
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