Application of up-regulating agent of HR or NHEJ pathway in preparation of medicament for treating diabetes mellitus and preventing diabetic individual tumorigenesis
A technology of diabetes and diabetic blood vessels, which is applied in the field of biomedicine and can solve the problems of unsatisfactory clinical effects of diabetic complications
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[0090] 1. Method
[0091] 1. Viral vector construction for overexpressing EWS (named: E-OE) or Fbxw7 (named: F-OE) protein
[0092] (1) Amplify the nucleotide sequences of EWS and Fbxw7 by PCR reaction (the sequence of EWS is shown in SEQ ID NO: 1, and the sequence of Fbxw7 is shown in SEQ ID NO: 2), and the two ends are connected with EcoRI and BamHI restriction sites point;
[0093] (2) double restriction digestion of PCR products and purification;
[0094] (3) Also double digest the H201pAdeno-MCMV-EGFP-3FLAG vector plasmid (Shanghai Heyuan Biotechnology Co., Ltd.), recover and purify the large fragment, and insert the EWS or Fbxw7 gene fragment in step (2) using molecular cloning technology instead of EGFP to the EcoRI and BamHI restriction sites of the shuttle plasmid;
[0095] (4) Colony PCR screens transformants, and the screened positive clones are sequenced and verified;
[0096] (5) Sequencing verifies the correct clones, and extracts high-purity plasmids;
[00...
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