Dna Polymerase - Ligase Detection Cisplatin Metabolism Related Gene Snp Method To Establish

As the new genetic marker, single nucleotide polymorphism(SNP) has been extensively applied to the diagnosis and treatment of diseases, pharmacogenomic research, genetic map, medicolegal identification, plant breeding and so on due to it's widespread distribution, high genetic stability and easy to analysis.There are three million SNPs sites in human genome and SNP occur at a frequency of approximately 1 in 1000 base pairs throughout the genome. SNP are distributed throughout the genome and these variations are associated with diversity in the population and individual metabolism to medicine. According to the patient's genotype, we should adopt the individualized treatment. Antitumor agent cisplatin, for example, cut the nucleotide into fragments, unable the DNA to duplicate or transcript and ultimately lead the tumor cells to apoptosis. Cisplatin can be used for non small-cell lung cancer and other solid cancers. Inherited SNP differences between individuals appear to determine each patient's adverse drug reactions in the treatment. The scientists have discovered many genes which affect the treatment of cisplatin. There are 12 genes reported and the related SNP are more than 15. By analysis these SNP polymorphism, we will be able to design different treatment for each patient.For the research of SNP, the difficulty lies in establishing the effective detection methods. We have developed a new method-the DNA ligase combined DNA polymerase detection reaction on the basis of other detection methods. Established from the ligase detection reaction (LDR),we utilized the DNA polymerase which could enhance the pairing of the bases in the process. Our method divided into four parts. (1) Template amplification of SNP: This step will amplify the signal of SNP and improve the accuracy of the results. (2) Polymerization-ligase reaction:On the basis of DNA ligase enzymatic specificity, we utilize the specification of DNA polymerase and make the detection results more accurate. Because of only one couple of probes used in the detection of every SNP, the detection cost is reduced and the job is easier. (3) Amplification of general primer, P3 and P4:Applying the general primer of P3 and P4 make our detection to be generalization and it reduced the work loads. (4) Applying agarose gel electrophoresis or oligonucleotide gene chips:to detect the SNP polymorphism.We used the DNA polymerase-DNA ligase detection reaction and the agarose gel electrophoresis to detect the SNP of tissues samples of 10 cases of non small-cell lung cancer. The results showed that 1 case had two heterozygote of the SNP and 3 cases had a mutant SNP respectively. Aiming at these patients'individuality, we offered the corresponding guides for every patient with the information gathered.