Molecular cloning and confirmation of MSH2, an essential gene of human, involved in DNA mismatch repair

Sarita Kumari; C Rajesh

Molecular cloning and confirmation of MSH2, an essential gene of human, involved in DNA mismatch repair

Authors
Sarita Kumari, C Rajesh

Abstract
Mismatch repair (MMR) is one such pathway that repair mismatches generated during DNA replication. Errors in MMR cause certain types of cancer including hereditary non-polyposis colorectal cancer, genome instability, abnormalities in meiosis, sterility in mammalian systems and resistance to certain chemotherapeutic agents. In eukaryotes, MSH2 (MutS homologue 2) is a major player in MMR, acting in combination with MSH3 or MSH6 as a heterodimer. MSH2 has been involved in a variety of processes which protect the integrity of the genome. In this study hMSH2 gene was cloned to identifying their role and understanding the functional diversity of protein that will contribute towards the repair mechanisms. Using in frame designed primers through PCR strategy and cloned full-length cDNA fragment hMSH2 gene into pET32b expression vector. The sequence contained an open reading frame of 2805 bp coding for a putative protein of 935 amino acids. Recombinant C-3 clone (pET32b-hMSH2) observed as positive after restriction digestion with expected band size (~8.6kb). This recombinant clone of hMSH2 gene, further used to express the protein and purified protein used to understand the role of mismatch repair protein MSH2.

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