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Single-nucleotide polymorphisms (SNPs) in pre-miRNAs may alter microRNA (miRNA) expression levels or processing and contribute to susceptibility to a wide range of diseases. We investigated the correlation between four SNPs (rs11614913, rs3746444, rs2910164, and rs229283) in pre-miRNAs and the risk of asthma in 220 asthma patients and 540 controls using polymerase chain reaction-restriction fragment length polymorphism methodology and DNA-sequencing. There were significant differences in the genotype and allelic distribution of rs2910164G/C and rs2292832C/T polymorphisms among cases and controls. The CC genotype and C allele of rs2910164G/C were significantly associated with a decreased risk of asthma (CC vs. GG, odds ratio [OR] = 0.51, 95% confidence interval [CI]: 0.31-0.82; C vs. G, OR = 0.74, 95% CI: 0.59-0.93). Similarly, the TT genotype and T allele of rs2292832C/T were significantly associated with a decreased risk of asthma (TT vs. CC, OR = 0.56, 95% CI: 0.33-0.95; T vs. C, OR = 0.71, 95% CI: 0.53-0.95). However, no significant association between the other two polymorphisms (i.e., rs11614913C/T and rs3746444C/T) and the risk of asthma was observed. Our data indicate that rs2910164G/C and rs2292832C/T may play a role in the development of asthma. [ABSTRACT FROM AUTHOR]

Investigates the haplotype frequencies of Y-chromosome STR loci in Chinese population. Development of a multiplex amplified for these Y-STR markers; Sufficiency of the sample size for the research purpose; Sensitivity and reliability of using non-denaturing polyacrylamide gel electrophoresis and visualized silver staining in the analysis of the PCR products.

Objective: To obtain data of polymorphism distribution of 10 short tandem repeat (STR) D1S2145, D3S2433, D5S1507, D5S2502, D8S2319, D9S926, D16S767, D17S2181, GATA140E03, GATA196B10 in Chinese Han population in Chengdu and to evaluate their usefulness in the field of forensic science and their species specificity., Methods: DNA of 100 unrelated individuals of Chengdu Han population was extracted with Chelex method, amplified by PCR, then typed with silver staining after polyacrylamide gel electrophoresis(PAGE). Ten different animals were selected as the controls in this study for evaluating the species specificity of the ten STR loci., Results: In the ten STR loci of Chengdu Han population, 6, 5, 8, 5, 6, 7, 7, 5, 7 and 7 alleles were found, respectively. 17, 14, 28, 15, 16, 18, 15, 14, 19 and 21 genotypes were observed in the ten loci, respectively. The allele and genotype frequency distributions of the ten loci were detected no deviation from the Hardy-Weinberg law of equilibrium. By comparison with the data from 10 different animals, the species specificity of D3S2433, D5S1507, D5S2502, D8S2319 and GATA196B10 was good, but part of animals had amplification product at typing field of the other loci., Conclusion: The 10 STR loci mentioned above are highly polymorphic and can be used in the forensic personal identification and paternity testing.

Objective: To establish a fluorescent multiple amplification system of 16S rRNA and Cytb genes located in mitochondrial DNA for species identification., Methods: A pair of primers of 16S rRNA gene and Cytb gene of the mitochondrial DNA was designed with the software Primer 5.0 to construct a multiple amplification system. The amplified products from human and five species of animals, including cattle, pig, dog, chicken and grass carp were analyzed by 310 Genetic Analyzer., Results: The amplified products of these samples showed two peaks. The common one was 358bp and the specific one different in unique species was between 231bp and 256bp., Conclusion: The multiplex amplification system can exactly distinguish the species of human from five common animals.

Objective: To evaluate the forensic utility of Y-single nucleotide polymorphisms (SNPs) markers., Methods: Allele-specific PCR, restriction enzyme digestion or direct PCR were performed to examine 10 different SNP loci on Y chromosome, namely M9, M15, M45, M89, M95, M122, M134, M145, M173 and P25 in 161 Chinese Han males., Results: A total of 8 of the 10 SNPs are reported to be polymorphic in Chinese. The gene diversity for the loci showing polymorphism ranged from 0.988/0.012-0.752/0.248, with a power of discrimination 0.094-0.373. Loci M122 and M134 were the most polymorphic markers in Chinese Hans. Nine different haplogroups with frequencies from 1.2% to 51.6% were observed and 3 of the haplogroups-K*(x O2a, O3, P), O3*(x O3e) and O3e were found in 75.2% of Chinese Hans., Conclusion: A comprehensive gene diversity data of Y chromosome and haplogroups were obtained in Sichuan Han population, which will be served as the base for using these Y-SNP markers in forensic medicine and individual identification in Sichuan Hans.

Objective: To formulate recommendations in the evaluation of results of genetic analyses in paternity testing under considering mutations., Methods: A total of 15 short tandem repeat(STR) loci were employed for this study, which were included CSF1PO, FGA, TH01, TPOX, VWA, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D21S11, PentaD and PentaE. Both 100 cases of true trio and 100 cases of false trio were investigated., Results: The numbers of mismatch alleles in different STR loci were observed in 100 cases of false trio. The different distributions of paternity index were obtained, including the changes of paternity index in each case of true trio under simulated mutations., Conclusion: In order to avoid the effect of mutations, the exclusion of paternity was never considered on the basis of a single locus. The threshold values of the combined probability of exclusion and the paternity index were important for both exclusion and inclusion of paternity. The scientific evidence for paternity testing can be obtained when both the combined probability of exclusion and the paternity index meet the threshold values. However, when either the combined probability of exclusion or the paternity index can not meet the threshold values, more genetic markers should be added.

Objective: To develop a multiplexed mutagenically separated PCR (MS-PCR) for single nucleotide polymorphism (SNP) loci typing in mitochondrial DNA coding regions and to study the applications in investigating the allele frequencies and haplotypes of four SNP loci in mitochondrial DNA coding regions in Chinese Chengdu Han population., Methods: Four SNP loci C12705T, A8701G, G8584A and C10400T, two allele specific forward primer with 4 bases different in size and a common reverse primer were designed for SNP typing. The primers simultaneously were amplified in a single tube. The genotyping of SNPs was determined by the two allele specific fragments different in size after polyacrylamide gel and silver staining., Results: The different SNP loci comprised a single band with different size respectively. Typing results were completely consistent with those by direct sequencing. The allelic frequencies of C12705T, A8701G, G8584A and C10400T were 0.3813/0.6187, 0.4813/0.5187, 0.8250/0.1750 and 0.4938/0.5062 respectively. A total of 6 different haplotypes was identified and the genetic diversity reached 0.7137., Conclusion: Multiplexed MS-PCR is a simple, rapid, accurate and efficient method for SNP typing, which will be very powerful for SNPs in the database establishing of mitochondrial DNA coding regions, the testing of forensic and population genetics research.

Objective: To obtain the data in polymorphism distribution of the five short tandem repeat (STR) loci: D18S979, D11S2014, D18S548, D1S1667 and GATA164F07 of Chinese Han population in Chengdu, and to evaluate their usefulness in the field of species specificity in forensic science., Methods: PCR, polyacrylamide gel electrophoresis (PAGE) and silver staining techniques were used to analyze the DNA samples from 100 unrelated individuals of Chinese Han ethnic group in Chengdu. Twelve different animals: monkey, pig, dog, bull, goat, chicken, duck, eel, mudfish, rabbit, guinea pig and mouse were selected as controls in this study for evaluating the species specificity of the five STR loci., Results: Six alleles and twelve genotypes were observed in D18S979. Five alleles and eleven genotypes were observed in D11S2014. Five alleles and thirteen genotypes were observed in D18S548. Seven alleles and nineteen genotypes were observed in D1S1667. Six alleles and fourteen genotypes were observed in GATA164F07. The genotype distributions of the five loci were analyzed by some related software and no deviation from the Hardy-Weinberg equilibrium was observed. Evaluated by way of using different animals as controls, monkey had amplification products at the extra-typing field of D18S979, D11S2014 and D1S1667. Bull, dog and eel had amplification product at typing field of D18S979, and pig, duck, mouse and rabbit had weak product. Bull had weak product at the typing field of D18S548. Dog, goat and eel had product at the typing field of D1S1667. Dog had weak product at the typing field of GATA164F07. Mudfish, chicken and guinea pig had no amplification product at the five loci., Conclusion: These data indicate that D18S979, D18S548, D1S1667 and GATA164F07 are highly polymorphic and D11S2014, D18S548 and GATA164F07 can play a key role in species identification.

Objective: In order to increase significantly the discriminatory potential of Y-STR systems available to the forensic community, we have developed a system capable of simultaneously amplifying 9 Y-STR loci by fluorescence-labeled multiplex PCR technique., Methods: Primers of STR loci DYS434, GATA-A10, DYS438 and DYS439 were labeled with 6-FAM, primers of STR loci DYS531, DYS557, DYS448 were labeled with HEX, and primers of STR loci DYS456, DYS444 were labeled with TAMRA, respectively. PCR products were analyzed using capillary electrophoresis and GeneScan Software on the ABI Prism310 DNA Analyzer. Series experiments were carried out to evaluate the useful value in forensic application such as the sensitivity, male specificity and genotyping DNA different tissues of the same individual., Results: 9 Y-STR loci were exactly determined following optimization of the polymerase chain reaction. In a sample of 120 males, a total of 105 different haplotypes was identified, 97 of them being unique. Overall, haplotype diversity was 0.996 8. It was proved that genotyping of these 9 Y-STR loci in some sexual crime should be prior to that of automosomal STR., Conclusion: The results suggest that the newly constructed 9-plex will be very powerful for establishing Y-STR database, population genetic studies and mixture stains identification.

Objective: To obtain the data in polymorphism distribution of the seven short tandem repeat (STR) loci: D1S2142, D1S3733, D2S1774, D3S2459, D21S1409, D21S1437 and D21S2055 of Chinese Han population in Chengdu, and evaluate the polymorphism data usefulness to the forensic science., Methods: PCR, polyacrylamide gel electrophoresis (PAGE) and silver staining techniques were used to analyze the DNA samples from unrelated individuals of Chinese Han ethnic group in Chengdu., Results: Eleven alleles and twenty-three genotypes were observed in D1S2142. Eight alleles and nineteen genotypes were observed in D1S3733. Eight alleles and fifteen genotypes were observed in D2S1774. Seven alleles and nineteen genotypes were observed in D3S2459. Six alleles and twelve genotypes were observed in D21S1409. Nine alleles and twenty-six genotypes were observed in D21S1437. Twenty alleles and seventy-seven genotypes were observed in D21S2055. The genotype distributions of the seven STR loci showed no deviation from the Hardy-Weinberg equilibrium. The parentage testing of 50 cases revealed an autosomal codominant inheritances and no mutations happened to seven STR loci., Conclusion: These data indicate that D1S2142, D1S3733, D2S1774, D3S2459, D21S1409, D21S1437 and D21S2055 have good polymorphism, with high probability of exclusion and probability of discrimination power as well as being loci available as the candidate genetic markers to forensic parentage testing and personal identification.

Objective: To construct STR slippage model and study factors involved in this procedure., Methods: DNA samples were amplified with the technology of Degenerate oligonucleotide- primed PCR, then their products were taken as later DNA template and their STR genotype were analyzed by optimizing several factors., Results: STR slippage model was constructed., Conclusion: Several factors were involved in the produce of STR slippage, such as amount of modulate DNA, concentration of MgCl2, property of DNA polymerase, motif sequence of STR loci, sample, etc.

Objective: To understand the role of mitochondria associated signaling pathway in the apoptosis of human vascular endothelial cell induced by homocysteine (Hcy)., Methods: The mRNA and protein expression levels of the up-stream signaling molecules of caspase 3, Bcl 2, caspase 9, and cytosolic cytochrome-c, were investigated. The in vitro cultured human umbilical vein endothelial cells with homocysteine at different concentrations were incubated for 24 h. The expressions of Bcl 2 and caspase 9 at mRNA and protein levels were analyzed by reverse transcription-polymerase chain reaction(RT-PCR) and Western blot. Cytochrome-c in cytoplasm was also detected by Western blot., Results: The expression levels of three signaling molecules were all down-regulated by homocysteine at both mRNA and protein levels in a dose-dependent manner., Conclusion: Homocysteine could affect the formation of apoptosome through repressing the expression of Bcl 2 gene and release of cytochrome-c from mitochondria. Decreasing of apoptosome could disturb the activation of caspase 9. The results also indicate that the mitochondria pathway is not the major signaling pathway involved in Hcy-induced apoptosis.

Objective: To build the four STR loci typing system by fluorescence labeled Multiplex-PCR technique, applied in the parentage test and personal identification in forensic medicine., Methods: The primer of D3S1754 and D1S549 were labeled with 6-FAM and TMR respectively, primers of D4S2366 and D12S375 were labeled with HEX. Multiplex-PCR products were analysed on the ABI PRISM 310 Genetic Analyzer where the Data Collection Software 3.0, the GeneScan Analysis Software 3.7NT and the Genotyper 3.7NT Software were used. This typing system has been emploied in the parentage test and personal identification of casework., Results: A method of typing four STR loci by fluorescence labeled Multiplex-PCR technique had been constructed. It has showed good sensitive and stability, and met the needs of parentage test and personal identification in forensic medicine., Conclusion: The constructed method can be used in studying genetic polymorphisms and parentage test or personal identification in forensic medicine.

To understanding the allele structure and genetic polymorphisms at five STR loci in Chinese Han population, and construct a preliminary database, EDTA-blood specimens were collected from unrelated individuals. DNA samples were extracted with Chelex method and were amplified by PCR technique. The PCR products were analyzed using both PAGE horizontal electrophoresis with discontinuous buffer system and automated fluorescence detection approach. As a result, three STRs consist of simple repeat motifs, while two STR contain a complex repeat structure. The STR polymorphisms at all of the five loci have been observed in Chinese Han population. In a word, the obtained data are beneficial to understanding the population genetics of the five STR loci in Chinese Han population. As a simple approach, the PAGE horizontal electrophoresis can be employed for typing the five STR markers.

Objective: To solve the problems in the accuracy and standardization of short tandem repeats-polymerase chain reaction (STR-PCR) typing, the authors adopted the molecular clone technology in producing the standard allelic ladders of D1S1676, D2S2735, D11S1977 and D22S444 loci and applied them in a population study on the Hans in Chengdu, China., Methods: PCR was used to produce several different allelic fragments of these loci. PCR products were eluted from the gel and re-amplified by PCR. The purified allelic fragments were then blunt-end subcloned individually into the pGEMR-T plasmid vectors and the recombinant were transfected into competent E.coli DH5alpha TM cells. The results of sequencing confirmed that the size and the construction of the inserts were correct. The recombinant plasmids DNA with the inserts were then used as template for re-amplification to generate the four loci standard ladders., Results: The authors succeeded in producing large quantity of standard allelic ladder of these four loci, with which the genetic polymorphisms of these loci in Chengdu Han population of China were studied., Conclusion: This method is of high value for forensic DNA typing to construct standard ladders. D1S1676, D2S2735 loci are robust for forensic analysis in Chinese Han population, whereas the value of D11S1977 and D22S444 loci is limited.

Objective: We focus on developing a multiplex PCR system for Y-STR loci that can be detected by double fluorescent system and assessing their usefulness in forensic mixture samples., Methods: The primers of four Y-STR loci (DYS-GATA-A10, DYS531, DYS557 and DYS448) amplified by multiplex PCR technique were labeled with fluorescence, then the PCR products of these Y-STRs loci were detecting and typing by ABI PRISM310 Genetic Analyzer., Results: When 120 unrelated individuals from the Han population in Chengdu were detected by the system, Y-GATA-A10, DYS531, DYS557 and DYS448 showed 5, 5, 8, 7 alleles, respectively. A total of 78 different haplotypes was identified and the genetic diversity reached 0.9881. To the three cases of mixture stains failed by using conventional autosomal STR analysis, our multiplex system drew conforming conclusion comparing to the suspect's Y-STRs genotypes., Conclusion: Our results show that the multiplex system of four Y-STR will be very powerful for Y-STR database establishing, the paternity testing and mixture stains identifying.

Objective: To develop a set of new markers for forensic application, the authors have chosen 6 short tandem repeat(STR) loci to study the allele frequencies and species specificity in Chinese Han population in Chengdu., Methods: One hundred and ten EDTA-blood samples were collected from the unrelated individuals in Chengdu city, Sichuan province. DNA was extracted by Chelex-100 and amplified by the polymerase chain reaction(PCR). Polyacrylamide gel electrophoresis (PAGE) and silver staining were used to analyze the PCR products., Results: The polymorphisms of all 6 STR loci have been obtained in Chinese Han population in Chengdu, the alleles of D4S2366, D4S2367, D6S474, D6S1281, D2S1396 and D20S601 being 7, 7, 6, 7, 5, 7, the observed heterozygosity of them being 0.802, 0.708, 0.770, 0.627, 0.542, 0.672, the discrimination power of them being 0.887, 0.828, 0.849, 0.848, 0.794, 0.865; and the power of exclusion of them being 0.602, 0.441, 0.544, 0.325, 0.227, 0.386. Evaluated by comparison with the data from 14 different animals as controls, the 6 STR loci contain good specificity of human beings., Conclusion: The 6 STR loci are highly polymorphic and can play a key role in species identification. They are new candidate markers for forensic personal identification and paternity testing.

Objective: To obtain the data of polymorphism distribution of the two STR loci D2S2944, D1S2134 in Chinese Han population in Chengdu and evaluate their usefulness in the field of forensic science., Methods: PCR, polyacrylamide gel electrophoresis (PAGE) and silver staining were used to analyze 120 unrelated individuals of Chinese Han ethnic group in Chengdu. Fourteen different animals: monkey, pig, dog, bull, goat, chicken, duck, fish, cat, rabbit, Guinea pig, mouse, eel and frog were selected as controls in this study for evaluating the species specificity of the two STR loci., Results: Eight alleles and twenty-two genotypes were found in D2S2944. The observed heterozygosity (h), discrimination power (DP), polymorphism information content (PIC), chance of paternity exclusion power (EP) were 0.824, 0.925, 0.78, 0.644 respectively. Ten alleles and twenty-six genotypes were observed in D1S2134. The h, DP, PIC and EP were 0.769, 0.920, 0.79, 0.543 respectively. The genotype distributions of the two loci were analyzed by some related software and no deviation from the Hardy-Weinberg equilibrium was observed. Evaluated by way of using different animals as controls, the two STR loci of human beings were found to have good specificity., Conclusion: These data indicate that D1S2134 and D2S2944 are of good polymorphism, high EP and DP, and can be applied as the candidate genetic marks to forensic parentage testing and identification. The methods to analyze them are simple, dependable and repeatable.

Objective: To investigate the association between a -344 C/T polymorphism of aldosterone synthase gene (CYP11B2) and the left ventricular structure and function of patients with dilated cardiomyopathy (DCM) in China., Methods: Eighty-nine patients with DCM were genotyped by PCR-RFLP, and were divided into TT, TC, CC and TT+CC four groups by their genotypes, respectively. Left ventricular structure and function were assessed with echocardiography., Results: There was no significant difference between four groups in left ventricular diameters, mass and ejection fraction in patients with DCM., Conclusion: The data were not in favor of a significant association between the aldosterone synthase gene CYP11B2 and the left ventricular structure and function in patients with DCM.

Objective: To establish a simplified method of multiplex PCR based on chimeric primers for STR loci., Methods: A set of chimeric primers and universal primers were designed to carry out a multiplex PCR for amplifying four short tandem repeats(STR) loci. Then the amplified STR loci were detected by polyacrylamide gel electrophoresis and the gels were sliverstained., Results: A quadriplex STR system was developed on the basis of both chimeric and universal primers., Conclusion: The STR multiplex PCR based on both chimeric and universal primers hss been achieved readily and reproducibly by simple adjustment of the individual primer concentrations. The use of chimeric primers provides a method for primer design that eliminates the multiple optimization steps involved in developing the multiplex PCR.

To investigate the genetic polymorphisms of three short tandem repeats loci of chromosome X in Chinese Han population in Chengdu area and its use in forensic science. Three X-chromosome linked short tandem repeat loci were analyzed by PCR followed by polyacrylamide gel electrophoresis. Hardy-Weinberg equilibrium was tested and forensic interested value was calculated . The power of exclution of DXS6804, DXS9896 and GATA144D04 is 0.5990, 0.6220, 0.4280, respectively. The result showed that all the three STR loci were polymorphic among 100 unrelated females and 120 unrelated males from Chinese Han population. Chi(2) tests demonstrated that genotype frequencies in females did not depart from Hardy-Weinberg equilibrium. Three X-chromosome linked short tandem repeat loci have high polymorphism, they can be applied to forensic medicine and population genetics.

Objective: To understand the allele structure and genetic polymorphism at three STR loci on chromosome X in Chinese Han population and make an evalution of their forensic application., Methods: EDTA-blood samples were collected from the unrelated individuals in Chengdu, China. After being extracted with Chelex method, the DNA samples were amplified by PCR technique. The PCR products were analyzed by PAG electrophoresis and the approach of automated fluorescence detection. Hardy-Weinberg equilibrium of females was tested and every forensic interested value was calculated., Results: The polymorphisms of all 3 STR loci were obtained from 100 unrelated females and 120 unrelated males from Chinese Han ethnic group. Chi-square tests on the genotype frequencies in females did not reveal deviations from Hardy-Weinberg equilibrium., Conclusion: The obtained data are beneficial to understanding the population genetics of the three STR loci in Chinese Han population. For forensic genetics, the obtained data can be used to calculate the probabilities dealing with the paternity test and the individual identification.

Objective: In Caucasian population, the most common molecular basis for C8 beta deficiency s a single C to T transition in exon 9 of C8 beta gene resulting in a stop codon. In previous family studies, two individuals were identified with C8 beta complete deficiency and were found to be only heterozygous for this mutation. This study was conducted by the present authors in search of other possible causes for these two C8 beta deficient individuals., Methods: Using direct DNA sequence analysis of all exon-specific PCR products of the C8 beta gene from these two C8 beta deficient patients and their descendants., Results: Two other C to T transitions at base 298 and 388 in exon 3 were detected, which could also create a termination codon. The descendants from one of the deficient patients were also analysed for the mutations, and it could be demonstrated that the two C to T mutations in exons 9 and 3 are segregating independently., Conclusion: These two mutations, which create a termination codon, are sufficient to explain the complete C8 beta deficiency in both patients.

To understand the genetic polymorphism of 10 STR loci on chromosome 9 and reveal the difference on the linkage maps of different populations, the author used a set of nine tetranucleotide- and one tritranucleotide-repeat STR markers on chromosome 9 chosen from Genome Data Bank. EDTA-blood was collected from unrelated family in Baiyin City, Gansu province. DNA was extracted by Chelex-100 method and amplified by the polymerase chain reaction (PCR). The PCR products were analyzed using PAGE electrophoresis. We calculated the allele frequencies and heterozygosity and PIC of every ten STR loci. Using the data of Chinese Han pedigree, we constructed a genetic map of chromosome 9 making up with 10 STR loci. Then we compared our map with the map in GDB. Our results indicate that there are some differences on the genetic map between Chinese and GDB. That means, it is essential to do some population genetic evaluations before using the data of GDB into Chinese population.