Detection Of H19Imprinted Genetic Polymorphism From Chelex-100DNA Template And Its Distribution In Shanxi Population

Objective(1)We have revealed the H19imprinting (variable number tandem repeat) VNTR polymorphism using Chelex-100DNA as HhaI subtract in order to provide a non-time-consuming and easily manipulated approach for imprinting polymorphism detection in forensic casework.(2)We also investigated the human H19imprinting VNTR genetic polymorphism in Shanxi Han population to evaluate the significance of this polymorphism in forensic application.Methods(1) To optimize the amount of genomic DNA for H19imprinting VNTR detection, we chose phenol/chloroform extracting DNA with amount from15ng to30ng as substrate to reveal the imprinting VNTR polymorphism. The optimized genomic DNAs were divided into two group, control and experiment. The experiment group was processed HhaI digestion followed by PCR, while control group without HhaI digestion. The PCR products were visualized by agarose gel electrophoresis and EB staining To discriminate the parental profile the hetereozygous bands were scanned and analyzed by SPSS software.(2) One hundred blood samples from unrelated healthy individual were investigated to probe the genetic information on H19imprinting VNTR locus.Results(1) The optimized amount of substrate DNA for Hhal is15ng under the following condition:total volume of20μl and HhaI (NEB)30U at37℃for15 hours. In this optimized condition the maternal bands are nearly invisible on naked-eye for HhaI digestion from phenol/chloroform extracting DNA. While for HhaI digested Chelex-100DNAthe very faint bands originated from maternity can still be seen. Therefore we scanned the bands for either from paternity or maternity and analyzed them by SPPS17.0software. The statistical results showed that there are significant difference of optical density from paternity or maternity for paired T test.(P<0.05).(2) Five alleles in human H19imprinting VNTR locus were detected in100unrelated healthy individuals named6,7,9,10and11accordingly. The allele frequency distribution is consistency of Hardy-Weinberg equilibrium(P>0.05).For H19imprinting VNRT polymorphism the heterozygosity (h), the polymorphism information content(PIC) and probability of discrimination(DP) are0.74,0.621and0.892respectively.Conclusion(1) The human H19imprinting VNTR polymorphism could be detected from Chelex-100DNA template. By this way the H19imprinting VNTR polymorphism can be easily analyzed without time consuming phenol/chloroform extracting procedure.(2) The human H19imprinting VNTR polymorphism in Shanxi Han population belongs to high personal discriminating power polymorphism and can be used for forensic individual identification and parentage testing.