Isolation and Characterization of ABCB5 Gene Promoter Region from Zebrafish

Auni Haziqah, Sama Jamal (2018) Isolation and Characterization of ABCB5 Gene Promoter Region from Zebrafish. [Final Year Project Report] (Unpublished)

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Abstract

ABCB5 (ATP-binding cassette, subfamily B, member 5) protein consists of P-glycoprotein that affects the levels of plasma and intracellular drugs concentration that control therapeutic response. This study aims to isolate and characterize the promoter region of the ABCB5 gene from zebrafish (Danio rerio). Initially, DNA extraction was performed on zebrafish by using lysis buffer. Then, the primer for the ABCB5 promoter was designed and the gradient PCR was done with the optimum temperature 53.5°C to produce single amplicon with the approximate size of 1,500 bp. The amplicon was cloned into pGEM-T Easy Vector and transformed into Escherichia coli XLl Blue Competent cells with the transformation efficiency of 0.05x10° transform ants per µg. Colony PCR was conducted to determine the positive colony with promoter insert. Next, plasmid miniprep was conducted before sending the plasmid for sequencing. However, the BLASTn results showed that the sequence cloned is not ABCB5 promoter region from zebrafish. Hence, it is further suggested that cloning should be continued in order to determine the transcription factor which regulates ABCB5 in order for us to further und

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