Novel STR Loci On Human Chromosome 6q24-25.1 And Application Of Forensic And LOH Research In The Gastric Cancer

Background & ObjectiveSTR is short tandem repeats with rich polymorphism which exists in the human genome widely,also named microsatellite DNA. As the second generation of genetic marker,it has been widely applied in the area of personal identification, paternity testing, genetic mapping, linkage disequilibrium analysis,loss of heterozygosity of tumor suppressor genes,organ transplantation and so on. STR analysis greatly widens the area of forensic medicine and the study of STRP in populations is the foundation of its forensic application. A tumor suppressor gene is often inactivated by a two-hit mechanism that involves mutation of one allele and loss of heterozygosity (LOH) of the other.Loss of heterozygosity is a common genetic alteration found throughout the genome in most solid neoplasms. Regions of nonrandom LOH can indicate the presence of genes whose loss promotes neoplastic progression and may have prognostic significance.We aim to explore novels STR loci on human Chromosome 6q24-25.1 in odrer to develop a set of new genetic mkarers for forensic science,oncology phymatology and so on.To investigate the genetic polymorphism of new STR loci in Han population and evaluate the value of utility of them.Meanwhile,we analyzed LOH on 30 paired-sample patients using 4 STR markers distributed on 6q24-25.1,for further searching molecular mechanisms for the multiple pathogenesis of gastric cancer.MethodsThe DNA sequence of human chromosome 6q24-25.1 was browsed to find out novel STR loci with tetranucleotide repeats units. Primers for PCR were desinged using Primer3 software in the flank sequences. The new STR loci were analyzed with touch-down PCR amplification and gel electrophresis. We optimized its amplification conditions and searched for different alleles in the population and constructed the allele ladders of the novel STR in house.Allele frequencies of the novel STR loci were investigated in Han population of Henan province. The results were analyzed by PowerStats vl.2 and obtained the Population genetics data.The forensic parameters such as discrimination power(DP), probability of exclusion(PE), polymorphism information content(PIC), heterozygosity(H), allele ffequency, Matching probability (PM)and so on were calculated.Tumor and normal DNA were screened for PCR analysis using 4 STR markers mapping the areas between 6q24.1 and 6q24.2:D6SZ2,D6SZ7,D6SZ8,D6SZ9,which was to detect LOH occurrence.ResultsWe found out 14 novel STR loci on 6q24-25.1 region,DP,H and PIC of 6 STR loci of which were higher and were also easy to be DNA testing accurately.They are nominated as D6SZ2,D6SZ6,D6SZ7,D6SZ8,D6SZ9,D6SZ12, respectively,the length of the products of which were 219bp,150bp,167bp,234bp,246bp,193bp respectively in the reference. These ten STR loci had 6,6,7,4,7,5 alleles in Han population of Henan province.Heterozygosity(H) were 0.555,0.555,0.585,0.712,0.667,0.750 respectively. The power of discrimination(PD) and the polymorphic information content(PIC) were0.857,0.811,0.821,0.803,0.829,0.827 and 0.65, 0.51,0.62,0.60,0.61,0.64 respectively.Of the 30 paired gastric cancer cases and normal tissues from the same individuals,LOH was observed in four cases.The frequency of LOH was the highest at marker D6SZ7(15.0%) on chromosome 6q24.2,followed by D6SZ2(11.1%) on chromosome 6q24.1, D6SZ8(9.09%) on chromosome 6q24.2, D6SZ9(6.25%) on chromosome 6q24.2.ConclusionUsing the DNAsequence information of 6q24-25.1 region,we found out 6 STR loci that have good genetic polymorphism and were easy to be tested accurately and then can be used in forensic DNA analysis and the study of human genetics.There are one tumor suppressor gene PLAGL1 at 6q24-25.1,which takes place LOH of the low frequency in the gastric cancer samples.