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A Python script to design primers for overlap extension PCR to ligate two DNA fragments

Ligating two or more DNA fragments is a regular operation for the subcloning and the engineering of vectors. The overlap extension PCR serves as a straightforward method to solve this issue. However, it takes a relatively long time to design the appropriate overlapping primers and the primers for th...

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Detalles Bibliográficos
Autores principales: Hu, Yunzhuo, Xu, Fei, Huang, Bifang, Chen, Xuanyang, Lin, Shiqiang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9632469/
https://www.ncbi.nlm.nih.gov/pubmed/36340189
http://dx.doi.org/10.7717/peerj.14283
Descripción
Sumario:Ligating two or more DNA fragments is a regular operation for the subcloning and the engineering of vectors. The overlap extension PCR serves as a straightforward method to solve this issue. However, it takes a relatively long time to design the appropriate overlapping primers and the primers for the full-length sequence, and there has not been a professional offline software for such kind of primer design. Here, we propose a Python script to search, calculate and sort thousands of combinations of primers for users according to the predefined parameters. The results of script running and experimental validation show that this script is capable of generating the optimal pairs of primers based on the proper melting temperatures and lengths of the primers, which facilitates gene modification in research.