Single primer-mediated circular polymerase chain reaction for hairpin DNA cloning and plasmid editing

Analytical Biochemistry
Jiansheng HuangNaishuo Zhu

Abstract

We developed and validated a universal polymerase chain reaction (PCR) method, single primer circular (SPC)-PCR, using single primer to simultaneously insert and amplify a short hairpin sequence into a vector with a high success rate. In this method, the hairpin structure is divided into two parts and fused into a vector by PCR. Then, a single primer is used to cyclize the chimera into a mature short hairpin RNA (shRNA) expression vector. It is not biased by loop length or palindromic structures. Six hairpin DNAs with short 4-nucleotide loops were successfully cloned. Moreover, SPC-PCR was also applied to plasmid editing within 3 h with a success rate higher than 95%.

References

Mar 23, 2002·Science·Thijn R BrummelkampReuven Agami
Mar 26, 2004·Nature·Patrick J PaddisonGregory J Hannon
Aug 12, 2004·Nucleic Acids Research·Lei ZhengJean-Louis Reymond
Jun 20, 2012·Journal of Visualized Experiments : JoVE·Daniel B StovallGuangchao Sui
Mar 17, 2015·Expert Opinion on Biological Therapy·Alesya A FokinaJean-Christophe François

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Citations

Mar 6, 2019·Antimicrobial Agents and Chemotherapy·Jiansheng HuangJiansong Ji
Dec 18, 2019·Antimicrobial Agents and Chemotherapy·Jiansheng HuangZhigang Zhao
Dec 16, 2018·Nature Communications·Amanda L MinogueSwathi Arur

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