Cloning of glucose oxidase gene in PKK233-3

Glucose oxidase [GO] has found a variety of industrial applications such as food, chemical and personal care industries. However one of the most important application of GO is used as diagnostic kits. The aim of study was isolation of GO gene from a recombinant vector [PET21aGO] and sub cloning and expression in PKK233-3 vector. Recombinant PET21a GO was extracted from E.coli DH5alpha and was digested with Restriction Enzymes; BamHI, Hindlll then isolated GO gene [1.8kb] and cloned in PKK233-3. PKK233-3GO was transformed in to E.coli DH5alpha. Our data demonstrated that the GO gene has expected size in agarose gel electrophoresis and also the cloned Go has a correct size after restriction analysis. The GO gene was cloned in prokaryotic host. This is a report of cloning of GO gene in Iran that can be used for further cloning of that gene in expression vectors for production of recombinant Enzyme

Study of XmnI polymorphism in 5 region of the apoAl gene in Iranian hyperlipidemic subjects

Several studies have demonstrated the relationship between polymorphisms in the ApoAl - CIH - AIV gene cluster and hyperlipidemia. This study was conducted to elucidate the association between polymorphism of ApoAI/XmnI and Iranian hyperlipidemic subjects. Total genomic DNA was prepared from seventy-six Iranian patient with primary hyperlipidemia and seventy-five normolipidemic subjects. The subjects in the control group were age-and sex-matched to the patients. Fragment of 392 bp for 5 region of the apoAl gene [C-2500T] was amplified by polymerase chain reaction [PCR]. In the hyperlipidemic group, the genotype frequency of X1X1, X1X2, X2X2 were 0.63, 0.24, 0.13, respectively. In the control group those were 0.81, 0.11 and 0.08, respectivley. There was a significant difference [p<0.05] between 2 groups. The rare allele [X2] was more frequent in hyperlipidemic group than in controls [p<0.01]. Various genotypes of apoAl/Xmnl had no significant effect on lipids or apoAI levels in hyperlipidemic group. The above results show that polymorphism ApoAl/XmnI is associated with hyperlipidemia in Iranian hyperlipidemic subjects. Therefore, our data confirmed the previously reported association between genetic polymorphism ApoAI/XmnI and hyperlipidemia

Isolation of Aspergillus Niger Glucose Oxidase gene and cloning of that prokaryotic host

Glucose oxidase [GO] is an enzyme, which use in food, chemical and personal care industries as well as glucose diagnostic kits. The aim of this study was PCR-mediated amplification of GO gene from Aspergillus niger genome and cloning of it in E. coli as a basis for production of recombinant GO in Iran. A. niger [2029] was cultured in media containing peptone, glucose and malt extract in 24'C for 48-72 h. Genomic DNA was extracted by sonication and freeze-thaw in liquid nitrogen in a lysis buffer containing EDTA and SDS. GO gene was amplified with designed primers under optimized PCR condition. The PCR product was cloned in pTZ57R plasmid using InsT/Aclone[TM] [Fermentas] and the constructed plasmid was transformed into E. coli DH5. Map of this plasmid was confirmed by restriction analysis and named pTZ57RGO. Our data showed the methods used in this study was adequate for culture and extraction of DNA from filamentous fungi such as A. niger. We showed amplified DNA fragment has expected size in agarose gel electrophoresis and also the cloned GO has a correct size after restriction analysis. The GO gene isolated successfully from A. niger via optimized PCR conditions and was cloned in prokaryotic host. This is the first report of isolation and cloning of GO gene in Iran that can be used for further cloning of that gene in expression vectors for production of recombinant enzyme