X染色体STR荧光复合扩增体系的建立及多态性

Objective To learn about the genetic diversity of X-STR and to collect data of population genetics, the six X-STR loci were tested. Methods DXS6801、 DXS9902、 DXS6809、 DXS6803、 DXS6804 and DXS6799 were amplified in a single PCR reaction. PCR products were analyzed using capillary electrophoresis and ABI prism 3100 Genetic Analyzer, with GeneMapper ID 3. 1 Analysis Software. Results Allele typing with the systems was successful for all of six loci. When 202 unrelated male and 62 unrelated female individuals from Guangdong Han population were tested, 8,6, 11, 10,6 and 9 alleles were detected for DXS6801, DXS9902, DXS6809, DXS6803, DXS6804 and DXS6799, respectively. Polymorphism information content is 0. 662 3～0. 837 6. Power of discrimination in females is 0. 824 2～0. 951 1. Mean exclusion chance for X-STR in standard trios with daughters is 0. 594 9～0. 817 4. Conclusion The six loci in the multiplex system provide high polymorphism information for forensic identification and paternity testing, particularly for difficult paternity deficiency cases     

X染色体STR荧光复合扩增体系的建立及多态性

目的 研究X染色体STR基因座的遗传多态性,建立群体遗传学数据.方法 利用荧光标记引物复合PCR技术,在同一反应管中同时检测 DXS6801、DXS9902、DXS6809、DXS6803、DXS6804 和DXS6799 6个X-STR 基因座,采用 ABI PRISM 3100 遗传分析仪电泳和 GeneMapper ID 3.1 软件进行基因分型.结果 用此体系同时分析6个 X-STR 基因座,检测264个广东汉族无关个体(男202人;女62人),DXS6801、DXS9902、DXS6809、DXS6803、DXS6804和DXS6799分别检出8、6、11、10、6和9个等位基因,多态性息含量(PIC)为0.662 3～0.837 6,女性个体识别率(PDF)为0.824 2～0.951 1,三联体非父排除率(MEC)为0.594 9～0.817 4.结论 本研究的6个X-STR基因座有高多态性和高个体识别率,在法医学个体识别和女孩的亲权鉴定中有重要应用价值.     

Investigator Argus X-12扩增体系在广东汉族人群的遗传多态性

目的 调查Investigator Argus X-12扩增体系的12个X染色体短串联重复序列(Xchromosome short tandem repeat,X-STR)基因座在广东汉族人群的遗传多态性.方法 采用荧光标记复合扩增和毛细管阵列电泳技术,对200名广东汉族无关个体(男性100名,女性100名)和103例家系样本(父-母-女三联体59例,母-子二联体44例)的DNA进行12个X-STR基因座分型.结果 在该群体中12个X-STR基因座共检出137个等位基因,其中包括9种稀有等位基因(off-ladder allele,OL allele).在162次减数分裂中共观察到6个突变事件.12个X-STR基因座的累积男性个体识别力为0.999 999 997,累积女性个体识别力为0.999 999 999,累积三联体非父排除率为0.999 999 988,累积二联体非父排除率为0.999 998 013.结论 Investigator Argus X-12系统在广东汉族人群中具有高度的多态性,对个体识别和亲权鉴定案件,尤其是特殊亲权鉴定案件极具应用价值.

Abstract：

Objective To investigate the genetic polymorphisms of 12 X chromosome short tandem repeat (X-STR) loci of Investigator Argus X-12 amplification kit in Guangdong Han population. Methods DNA samples from 200 unrelated individuals (100 males and 100 females) and 103 families (59 fathermother-daughter trios and 44 mother-son duos) were extracted and amplified with fluorescence labeled multiplex PCR system. PCR products were separated and genotyped with capillary array electrophoresis.Results One hundred and thirty-seven alleles,including 9 off-ladder alleles (OL allele) were observed at the 12 X-STR loci in the population. Six mutations were observed in 162 meioses. The combined power of discrimination (DP) was 0. 999 999 997 in males and 0. 999 999 999 in females, and the combined mean exclusion chance (MEC) was 0. 999 999 988 in the trio cases and 0. 999 998 013 in the duo cases. Conclusion Investigator Argus X-12 amplification system is highly polymorphic in Guangdong Han population and it is powerful for personal identification and paternity testing.     

Genetic polymorphism and relationship of 24 Y-STR loci among three ethnic minorities from GuizhouCSCD

Objective: To study the distribution of haplotypes of Y-chromosomal short tandem repeats (Y-STR) loci among three ethnic minorities from Guizhou, China. Methods: Twenty four Y-STR loci of 174 unrelated males were amplified with a Microreader™ 24Y Direct ID System kit. Capillary electrophoresis was carried out on an ABI 3100 Genetic Analyzer, and the data was analyzed with GeneMapper software. Results Seventy six haplotypes were identified for the 24 Y-STR loci among the three ethnic minorities, including 13 from the Qiangs, 35 from the Manchurians, and 28 from the Shes, with the corresponding Haplotype Diversity (HD) being 0. 7327, 0. 9578, and 0. 9344. Genetic distance between the Shes and Qiangs was relatively close, whilst that for Manchurians was relatively far. Conclusion: Analysis of the genetic characteristics and relationship of the three ethnic minorities from Guizhou can provide a reference for the study of their origin, evolution and patrilineal fusion. © 2018 MeDitorial Ltd. All rights reserved.     

EMILIN1基因多态性与蒙古族原发性高血压相关性研究

Objective To explore the relationship between genetic polyrnorphisms of 3 single nucleotide polymorphisms (SNPs) in the elastin microfibril interfacer 1 (EMILIN1)gene and essential hypertension. Methods A case-control study was conducted in which 201 hypertensive patients and 202 healthy controls in Mongolian population were enrolled, and the genotypes of rs3754734, rs2011616 and rs2304682 loci were analyzed using polymerase chain reaction-restriction fragment length polyrnorphism (PCR-RFLP) and direct sequencing techniques. Results There were significant differences in the frequencies of alleles and genotypes for the rs2304682 between the hypertensives and normotensives in the population(P＜0. 05). The frequency of the G-G haplotype established by rs3754734 and rs2304682 was significantly higher in the hypertensive patients (P＜0. 05). The frequencies of alleles and genotypes for the rs2304682 also had significant differences between the group with high diastolic blood pressure and normal diastolic blood pressure (P＜0.05). There were no significant differences in the frequencies of alleles and genotypes for the 3 SNPs between the group with high systolic blood pressure and normal systolic blood pressure (P＞0.05). Conclusion The rs2304682 locus in the EMILIN1 gene, as well as the haplotypes G-G constructed using rs3754734 and rs2304682, may associate with the susceptibility of essential hypertension in the Mongolian population. Also, rs2304682 may associate with the level of the diastolic blood pressure.     

EMILIN1基因多态性与蒙古族原发性高血压相关性研究

Objective To explore the relationship between genetic polyrnorphisms of 3 single nucleotide polymorphisms (SNPs) in the elastin microfibril interfacer 1 (EMILIN1)gene and essential hypertension. Methods A case-control study was conducted in which 201 hypertensive patients and 202 healthy controls in Mongolian population were enrolled, and the genotypes of rs3754734, rs2011616 and rs2304682 loci were analyzed using polymerase chain reaction-restriction fragment length polyrnorphism (PCR-RFLP) and direct sequencing techniques. Results There were significant differences in the frequencies of alleles and genotypes for the rs2304682 between the hypertensives and normotensives in the population(P＜0. 05). The frequency of the G-G haplotype established by rs3754734 and rs2304682 was significantly higher in the hypertensive patients (P＜0. 05). The frequencies of alleles and genotypes for the rs2304682 also had significant differences between the group with high diastolic blood pressure and normal diastolic blood pressure (P＜0.05). There were no significant differences in the frequencies of alleles and genotypes for the 3 SNPs between the group with high systolic blood pressure and normal systolic blood pressure (P＞0.05). Conclusion The rs2304682 locus in the EMILIN1 gene, as well as the haplotypes G-G constructed using rs3754734 and rs2304682, may associate with the susceptibility of essential hypertension in the Mongolian population. Also, rs2304682 may associate with the level of the diastolic blood pressure.     

Molecular screening of patients with nonsyndromic hearing loss from Nanjing city of China

Hearing loss is the most frequent sensory disorder involving a multitude of factors,and at least 50% of cases are due to genetic etiology.To further characterize the molecular etiology of hearing loss in the Chinese population,we recruited a total of 135 unrelated patients with nonsyndromic sensorineural hearing loss (NSHL) for mutational screening of GJB2,GJB3,GJB6,SLC26A4,SLC26A5 IVS2-2A>G and mitochondrial 12SrRNA,tRNA Ser(UCN) by PCR amplification and direct DNA sequencing.The carrier frequencies of deafness-causing mutations in these patients were 35.55% in GJB2,3.70% in GJB6,15.56% in SLC26A4 and 8.14% in mitochondrial 12SrRNA,respectively.The results indicate the necessity of genetic screening for mutations of these causative genes in Chinese population with nonsyndromic hearing loss.     

The distribution of DEN infected people in Dushan and Xingyi area of Yunnan-Guizhou Plateau, China

The dengue viruses （DEN, genus Flavivirus, family Flaviviridae） are mosquito borne and have caused 100 million cases of dengue fever each year in most tropical and subtropical areas of the world. However, in the Southwest area of Yunnan-Guizhou Plateau, China, the previous work demonstrated that different geographic strains of Aedes albopictus were susceptible to dengue virus. In this study, we collected 456 sera samples from patients with fever and 994 sera samples from healthy population in Dushan and Xingyi area of Yunnan-Guizhou Plateau, China. All sera samples were tested for dengue IgG by enzyme-linked immunosorbent assay （ELISA）. Patients＇ sera samples were tested for dengue IgM and DEN antigen was checked in the sera of 6 from 456 samples with which C6/36 cell in culated by IFA. The results indicate that these patients with fever were infected with DEN-2 and suggest that DEN infection had existed in Dushan and Xingyi area of Yunnan-Guizhou Plateau, China.     

DXS52(St14)在血友病甲基因诊断中的方法改进及应用

目的 改进DXS52(St14)在血友病甲(hemophilia A,HA)基因连锁分析中的实验方法并应用于基因诊断,报道DXS52位点与FⅧ基因发生重组的2个家系.方法 采用PCR和琼脂糖凝胶电泳对61个非倒位HA家系的DXS52位点进行基因检测,并用FⅧ基因内的Bcl Ⅰ、HindⅢ、Xba Ⅰ、STRl、STRl3、STR22和STR24这7个位点以及DXS52位点对这61个家系进行基因连锁分析.结果 DXS52位点在43个HA家系中可提供信息,可诊断率为70.5%(43/61).其中8个家系仅DXS52单个位点能提供信息,占13.1%;两个家系的DXS52与FⅧ基因发生基因重组.结论 采用新的实验条件可使DXS52位点基因检测得到准确清晰的实验结果.该位点可诊断率高,目前是HA连锁基因分析中不可缺少的诊断位点,但该位点位于FⅧ基因外,与FⅧ基因间存在重组可能,单独应用于诊断时应谨慎.

Abstract：

Objective To improve the experimental method of DXS52 (St14) and apply it to genetic testing for hemophilia A (HA). Methods PCR of DXS52 and agarose gel electrophoresis were performed for genetic testing in 61 non-inversion HA families. Linkage analysis of 7 loci within the FⅧ gene including Bcl Ⅰ , Hind Ⅲ, Xba Ⅰ , STR1, STR13, STR22 and STR24 were also carried out for the 61 families.Results DXS52 can provide information in 43 out of 61 families and the diagnostic rate was 70. 5%. Eight families can be diagnosed only by DXS52 locus, accounting for 13. 1%. Two families were found to have recombination between DXS52 and FⅧ. Conclusion The new experimental conditions can reach accurate and clear results in DXS52 genetic testing. This gene maker has high diagnostic rate, so it is an indispensable linkage analysis method in HA gene diagnosis. More caution should be paid when using the extragenic locus DXS52 to perform gene diagnosis because of its high recombinant rate with FⅧ.     

A sensitive method to detect the hepatitis B virus mutations by using solid phase PCR on oligonucleotide array

A sensitive method based on solid phase PCR on oligonucleotide array was established to detect two point mutations 1896G-A and 1901G-A in hepatitis B virus (HBV) DNA, in which 6 probes including these two point mutations were immobilized on modified glass slides through 5' terminal linker, while the 3' terminal was made to be free. The mutated loci were designed to locate on the last nucleotides of 3' terminal respectively, and the positive control probes lacked the last nucleotide of 3' terminal in comparison with the probes used for detection. Probes fixed on oligonucleotide array were also the solid phase amplification primers. One pair of liquid primers was used to amplify the short template product from whole HBV DNA. Using target DNA as template, the solid primers were extended under the action of Taq DNA polymerase and incorporated with Cy-3dCTP as marker. During the thermal cycling reaction, probes served as solid phase amplification primers and amplification products bound to the oligonucleotide array, which could be visualized by incorporation with fluorescent dyes. After amplification, the oligonucleotide array was washed, performed with laser scanning, and then used for quantitative analysis to obtain the information for mutations. The hybridization results were compared with DNA sequencing. It was demonstrated that in case of sample A, the ratios of fluorescence intensities in wide type and in the mutated types of 1896G-A and 1901G-A mutations in HBV were 3.81:1 and 1:3.79 respectively, while, in case of sample B, those were 1:2.89 and 1:3.03 respectively, indicating the presence of point mutations in these two loci. These results correlated to those obtained from DNA sequencing analysis in which the fluorescence intensity ratios in wide type and in the mutated types of 1996G-A and 1901D-A mutations in HBV were 1.26:1 and 1.67:1 respectively. From the above observations, it is evident that the method using solid phase PCR based on oligonucleotide array appears to be a sensitive and promising way to detect mutations with low-density.