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Shen C.-M.,Key Laboratory of Environment and Gene Related to Diseases | Shen C.-M.,Xi'an Jiaotong University | Zhu B.-F.,Key Laboratory of Environment and Gene Related to Diseases | Zhu B.-F.,Xi'an Jiaotong University | And 7 more authors.
Human Immunology | Year: 2010

The allele and haplotype frequencies of HLA-A, -B and -DRB1 loci in 10,000 healthy unrelated Han individuals living in the Guanzhong region of the Shaanxi Province were analyzed with the methods of SSO, SSP and SBT. Subsequently, these data were compared with results obtained in Han populations living in other regions as well as to other ethnic groups, using genetic distance measurements, neighbor-joining dendrograms and principal component analysis. In total 18 alleles were detected at the HLA-A locus, 46 alleles at the HLA-B locus and 14 alleles at the HLA-DRB1 locus. HLA-A02 was the most common HLA-A allele (29.70%), followed by A11 (18.70%), and A24 (15.75%); whereas at the HLA-B locus, the allele with the highest frequency was HLA-B13 (10.76 %), followed by B46 (7.93%), B51 (7.68%). At the HLA-DRB1 locus, the most common alleles were HLA-DRB115 (15.54%), DRB109 (13.18%) and DRB104 (11.21%). Three-loci haplotype analysis revealed that HLA A30- B13- DRB107 (4.11%), A02 -B46 -DRB109 (2.57%) and A33 -B58 -DRB117 (1.32%) were the most common haplotypes in this population. Four two-loci haplotypes, including HLA-A30-B43, A30-B53, B43-DRB107 and B73-DRB104 had significant linkage disequilibrium (relative linkage disequilibrium parameter equals to 1). Compared with other populations, our results indicated that the Han populations in different regions had a similar allelic diversity of HLA -A, -B, and -DRB1 loci. The Han population in the Guanzhong region of the Shaanxi Province had a close genetic relationship with the Northern and Southern Han populations. In summary, the similarities and differences of the HLA allelic diversity and haplotype structure between the Han population in the Guanzhong region and related populations, regarding HLA genotype distribution, provide basic information for further studies of the HLA heterogeneity and anthropological studies. © 2010 American Society for Histocompatibility and Immunogenetics.

Han Y.,Xi'an Jiaotong University | Han Y.,Key Laboratory of Environment and Gene Related to Diseases | Zhu W.-H.,Xi'an Jiaotong University | Zhu W.-H.,Key Laboratory of Environment and Gene Related to Diseases | And 10 more authors.
Journal of Xi'an Jiaotong University (Medical Sciences) | Year: 2010

Objective: To evaluate the feasibility of real-time quantitative PCR (RtPCR) with small volume regarding the stability, efficiency and reliability of amplification, and determine the optimal quantity of cDNA template suitable for small PCR volume. Methods: The experiment was carried out in 3 groups with 10, 15 and 20 μL reaction volume, respectively. In each group, rat β-actin mRNA was detected by RtPCR with 0.1, 0.2, 0.3or 0.4 μL rat cDNA as template, respectively. The amplification curve and melting curve were used to evaluate the reaction stability. The fitting of a curve of gradient templates against threshold cycle numbers was to show the reaction efficiency and the linear correlativity was to estimate the suitability of the template quantity. In addition, in order to estimate reliability, pristane-induced arthritis (PIA) rat model was established, and spleen TNF-α mRNA expression was detected by RtPCR with the selected reaction volumes. Results: The amplification of rat β-actin mRNA was specific and stable in 10 μL, 15 μL, and 20 μL PCR volume, and had a high efficiency. Furthermore, the standard curves fitted by 0.1-0.4 μL, gradient templates showed a significant linear correlation in each volume group. When the 10 μL and 20 μL. PCR volumes, and 0.2 μL, cDNA templates were chosen, the TNF-α mRNA expression in PIA rat spleen showed significant upregulation in both two volume groups as anticipated. Conclusion: The experiment shows that it is feasible in the RtPCR amplification to use the small reaction volume of 10 μL and 15 μL, which has good stability and reliability. And 0.1-0.4 μL templates are all suitable for the reaction system. PCR with small volume can not only save the reagents and template, especially rare clinical specimens, but also is helpful for the realization of high-throughput reaction.

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