Abdullah, Nur Azieyati
(2011)
Subcloning of cytochrome P450 2C19 (CYP2CL9)
In escherichia coli strain BL21.
Other.
Pusat Pengajian Sains Kesihatan, Universiti Sains Malaysia.
Abstract
Drug metabolism or biotransformation of drug undergo molecular alteration in which, it
converts nonpolar, lipohilic pharmacologically active drug molecules into polar,
inactive, or nontoxic metabolites in the body before excretion. In human, the
cytochrome P450 (CYP) enzymes are the major catalysts involved in the metabolism of
drugs and other xenobiotics that enter the body. The CYP2C19 enzyme is responsible
for the metabolism of several important drugs such as antidepressants. This study was
conducted to generate CYP2C19 enzyme recombinant for pharmocogenomics study.
Constructed eDNA of CYP2C19 was purchased from Origene, USA. The cloning
process was performed by using RapidShuttling™ Kit (Origene, USA). The CYP2Cl9
gene from TrueORF Entry Vector was transferred into an expression vector, pEX-CHis.
The desire plasmid was extracted using commercial QIAprep Spin Miniprep Kit
(Qiagen, Germany) and DNA sequence of CYP2C19 was confirmed by DNA
sequencing using ABI PRISMA ™ 3130x/ Genetyx Analyzer. The size of PCR product
was shown at -1.5kb on agarose gel. Desire DNA sequence of CYP2C 19 enzyme
shown same sequence as in Gene bank. As a conclusion, CYP2CJ9 gene was
successfully cloned. The combination of molecular biology and relative exposure to
CYP2C19 substrates and genetic testing eventually lead to a better appreciation to avoid
toxicological outcomes that may currently result from this enzyme and variations in its
expression.
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