SiteFinding-PCR: a simple and efficient PCR method for chromosome walking.
In this paper, we present a novel PCR method, termed SiteFinding-PCR, for gene or chromosome walking. The PCR was primed by a SiteFinder at a low temperature, and then the target molecules were amplified exponentially with gene-specific and SiteFinder primers, and screened out by another gene-specific primer and a vector primer. However, non-target molecules could not be amplified exponentially owing to the suppression effect of stem-loop structure and could not be screened out. This simple method proved to be efficient, reliable, inexpensive and time-saving, and may be suitable for the molecules for which gene-specific primers are available. More importantly, large DNA fragments can be obtained easily using this method. To demonstrate the feasibility and efficiency of SiteFinding-PCR, we employed this method to do chromosome walking and obtained 16 positive results from 17 samples.
Tan
Guihong
G
The National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University Beijing 100871, China.
Gao
Yin
Y
Shi
Miao
M
Zhang
Xinyue
X
He
Shanping
S
Chen
Zhangliang
Z
An
Chengcai
C
D017360:Arabidopsis / Q000235:genetics; D016386:Chromosome Walking / Q000379:methods*; D000458:Cyanobacteria / Q000821:virology; D017931:DNA Primers / Q000737:chemistry; D004269:DNA, Bacterial / Q000235:genetics; D016254:Mutagenesis, Insertional; D017900:Myoviridae / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D012231:Rhizobium / Q000235:genetics
D017931:DNA Primers; D004269:DNA, Bacterial