PubMed · 12013748
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Mohammad S Enayat. 2002. Restriction fragment length polymorphism.. https://doi.org/10.1385/1-59259-273-2%3A029
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We studied the distribution of Y-chromosome specific haplotypes (detected by the TaqI polymorphism of probes p49a,f) on a total of 328 Corsican males native of the regions of Ajaccio, Bastia and Corte. Three haplotypes are differential among regions: haplotype XV (A3 C1 D2 F1 I1), preponderant in the North of the island, haplotype V (A2 C0 D0 F1 I1) in the South, and haplotype XII (A3 C0 D1 F1 I0) in the highlands of the centre. Distribution of haplotypes can be explained by Corsican history and geography.
Hemophilia B is an X-linked, recessive disorder of hemostasis, caused by a defect in coagulation factor IX. To date, several restriction fragment length polymorphisms (RFLPs) have been identified within the gene for human factor IX. The incidence of these RFLPs differs significantly in different populations. In the present study, we analyzed the heterozygosity frequency and the allele frequency of three common intra- and extragenic polymorphic sites of the factor IX gene in Indian population. The main objective was to test the informativeness of two intragenic markers Dde I and Xmn I and one extragenic marker Hha I for carrier detection and prenatal diagnosis. The method used was polymerase chain reaction (PCR) and RFLP, which is economical yet simple to perform. In Indian population Dde I marker showed an informativeness of 69.0% followed by 38.0% for Hha I and 23.0% for Xmn I. The cumulative informativeness of these three markers was found to be 80 to 82%. A comparison of the heterozygosity rates of these three markers with the other ethnic groups showed that Indian population had almost similar pattern with the Caucasians and American blacks but differed significantly from the Orientals including Japanese, Chinese and Malays.
Polymerase chain reaction (PCR) is one of the most powerful tools in cloning genes. For the direct cloning of PCR products, T-vectors, which contain complementary 3'-thymidine overhangs, are widely used. In the present study, we developed a plasmid, pNB-T, which was constructed by cloning an XcmI cassette with a sufficient length of DNA (over 500 bp long) between two XcmI restriction sites into pBluescript SK(+). An XcmI cassette was made by nonspecific PCR using a primer containing recognition sequences of XcmI so that pNB-T can easily be converted into a T-vector by restriction of the plasmid with XcmI. In addition, the recognition sequences for BamHI and NcoI were added at 5'-end of the primer in order to facilitate subcloning of the gene cloned in the T-vector. The cloning efficiency of a PCR product, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene, was approximately 90%. Digestion of the recombinant plasmid containing the GAPDH gene with BamHI or NcoI liberated the DNA fragments with the expected size, demonstrating the usefulness of extra restriction sites. The method described in this report is quite simple and enables us to construct a variety of useful T-vectors.