The rRNA genetic locus is found in all prokaryotic organisms, and is highly conservative, although its relatively stable variations are found frequently in different bacteria. The utility of this locus as a taxonomic ...The rRNA genetic locus is found in all prokaryotic organisms, and is highly conservative, although its relatively stable variations are found frequently in different bacteria. The utility of this locus as a taxonomic and phylogenetic tool has been reported widely. This study, aimed at 16S rRNA gene (16S rDNA) and with the help of biomolecular methods, attempted to achieve the goal of rapid identification of common pathogens. In this study, 333 clinical isolated pathogenic bacteria were collected. Two pairs of primers were chosen and labeled with different fluorescent dyes and then used to amplify the genomic DNA extracted from bacteria. The PCR products were then detected by capillary electrophoresis-single strand conformation polymorphism (CE-SSCP). In order to pursue higher resolution and peak-separation effect, a high efficient separating medium, liner polyacrylamidedel (LPA), was put to use in this study. Finally, every bacteria colony generated distinct patterns from each other, which were easily to be used for identification. These results indicated that PCR-CE-SSCP was a rapid identification method for bacterial identification, with the aspects of high efficiency and high precision. Compared with traditional method, this technology is of great utility for clinical use especially for its high sensitivity.展开更多
AIM:To investigate the relationship between c.343A>G and c.2216A>C polymorphism sites in the CDH17 gene and colorectal carcinoma.METHODS:Ninety-three non-consanguineous colorectal carcinoma patients admitted to ...AIM:To investigate the relationship between c.343A>G and c.2216A>C polymorphism sites in the CDH17 gene and colorectal carcinoma.METHODS:Ninety-three non-consanguineous colorectal carcinoma patients admitted to the Department of Oncology at the First Affiliated Hospital of Zhengzhou University were included in this study.Ninety-three peripheral venous blood samples,of approximately one milliliter from each patient,were collected betweenDecember 2009 and August 2010.The genomic DNA of these peripheral venous blood samples were extracted and purified using a Fermentas Genomic DNA Purification Kit(Fermentas,CA) according to the manufacturer' s protocol.The single nucleotide polymorphisms(SNPs) of the liver-intestine cadherin(CDH17) gene c.343A>G and c.2216A>C were determined by the polymerase chain reaction-single strand conformation polymorphism method(PCR-SSCP) in 93 peripheral venous blood samples from patients suffering with colorectal carcinoma.Typical samples that showed different migration bands in SSCP were confirmed by sequencing.Directed DNA sequencing was used to check the correctness of the genotype results from the PCR-SSCP method.RESULTS:There was a significant association between the c.2216 A>C SNPs of the CDH17 gene and the tumor-node-metastasis(TNM) grade,as well as with lymph node status,in 93 peripheral venous blood samples from colorectal carcinoma patients.The genotype frequencies of A/C,A/A,and C/C were 12.90%,33.33% and 53.76%,respectively.There was a significant correlation between lymph node metastasis,TNM grade,and the genotype distribution(P < 0.05).The C/C genotype raised the risk of lymph node metastasis and the TNM grade.There was a significant difference in the TNM grade and lymph node metastasis between the A/A and C/C genotypes(P = 0.003 and P = 0.013,respectively).Patients with colorectal carcinoma carrying the C allele tended to have a higher risk of lymph node metastasis and have a higher TNM grade.The difference between the TNM grades,as well as the lymph node metastasis of the two alleles,was statistically significant(P < 0.01).CONCLUSION:The SNPs of the CDH17 gene c.2216 A>C might be clinically important in the prognosis of colorectal carcinoma.展开更多
By employing different gel components and electrophoresis conditions, the PCR-SSCP(polymerase chain reaction-single-stranded conformation polymorphism) method was used to detect the polymorphism(CCC or CGC ) of codon ...By employing different gel components and electrophoresis conditions, the PCR-SSCP(polymerase chain reaction-single-stranded conformation polymorphism) method was used to detect the polymorphism(CCC or CGC ) of codon 72 in exon 4 of p53 gene. The results展开更多
Objective: To study the relationship between the polymorphism of drug resistant gene rpoB and drug resistance against rifampicin(RFP) of M. tuberculosis L-forms, and to evaluate its clinical application. Methods: A to...Objective: To study the relationship between the polymorphism of drug resistant gene rpoB and drug resistance against rifampicin(RFP) of M. tuberculosis L-forms, and to evaluate its clinical application. Methods: A total of 52 clinical isolated strains of M. tuberculosis L-forms were collected. rpoB gene polymorphism was analyzed by polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) and conventional antimicrobial susceptibility test (AST). Their results were compared. Results: AST results showed that 38 of 52 clinical isolated strains were drug resistance (73.08％),while PCR-SSCP indicated 65.38％ (32/52) rpoB gene polymorphism. There was no statistic significance(χ2= 2.4914) between the 2 methods. Conclusion:Combined the application of PCR-SSCP with AST in detecting rpoB drug resistant gene polymorphism of M. tuberculosis L-form from pneumoconiosis patients with tuberculosis may have advantages at earlier diagnosis and guidance of clinical medications.展开更多
Objective To develop a high throughput mutational detection method by mutiple fluorescence-labeled polymerase chain reaction(PCR)products.Methods A total of 27 known mutations including 22 substitutions,3 insertions(1...Objective To develop a high throughput mutational detection method by mutiple fluorescence-labeled polymerase chain reaction(PCR)products.Methods A total of 27 known mutations including 22 substitutions,3 insertions(1,2 and 7 bp)and 2 deletions(1 and 2 bp)in the hepatocyte nuclear factor(HNF)-4α,glucokinase and HNF-1α genes were tested.During nested PCR,amplified fragments were labeled with three fluorescent dyes.PCR products were visualized with an ABI-377 fluorescence sequencer using 5% glycerol or 10% sucrose in nondenaturing gel conditions.Results Twenty-five of 27 variants(93%)could be detected by combining 5% glycerol and 10% sucrosegel matrix conditions.Twenty-two of 27(82%)and 18 of 27(67%)variants were identified using 5%glycerol and 10% sucrose conditions,respectively.Conclusion This fluorescence-based PCR single strand conformation polymorphism technique represents a simple,non-hazardous,time-saving and sensitive method for high throughput mutation detection.展开更多
BACKGROUND: Since single nucleotide polymorphisms (SNPs) can serve as gene markers, polymorphism profiles may help scientists to identify the full collection of genes that contribute to the development of complex dise...BACKGROUND: Since single nucleotide polymorphisms (SNPs) can serve as gene markers, polymorphism profiles may help scientists to identify the full collection of genes that contribute to the development of complex diseases such as cancer. The distribution of interleukin-10 (IL-10) promoter polymorphisms in Chinese Han ethnic patients with hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC) was investigated in this study. METHODS: The polymorphisms of IL-10 promoter region were detected by pulymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and sequencing. Sixty-six health controls, 42 patients with HBV infection, 30 HCC patients, and cell line SMMC-7721 were examined this way. RESULTS: Polyrnorphisms of T/C or T/N on-872 site occurred frequently in Han ethnic population. Pulyrnorphisms were detected in HBV and HCC patients and cell line SMMC-7721. The hotspot among the pulymorphisms was inserting base A between-1058 and-1057. CONCLUSION: Polymorphisms of IL-10 promoter in HBV and HCC patients may be associated with HBV infection and HCC development.展开更多
Poly （ADP-ribose） polymerase-1 （PARP-1） can exacerbate ischemic brain injury and lessen ischemic neuronal death, which may be associated with PARP-1 polymorphisms. The present study investigated human PARP-1 gene ...Poly （ADP-ribose） polymerase-1 （PARP-1） can exacerbate ischemic brain injury and lessen ischemic neuronal death, which may be associated with PARP-1 polymorphisms. The present study investigated human PARP-1 gene polymorphisms in various Chinese nationalities, the results of which could potentially help in the treatment and prevention of neurologic diseases. Genetic polymorphisms of seven exons in the PARP-1 gene, in 898 Chinese Han, Buyi, Shui, Miao, and Zhuang subjects, were investigated by PCR-single-strand conformation polymorphism. A single-strand conformation polymorphism variant in exons 12, 13, 16, and 17 of the PARP-1 gene was identified in 148 people, with two stationary bands showing three degenerative single strands. Results showed that the PARP-1 gene polymorphisms exist in various nationalities, and may act as a biomarker for susceptibility to disease.展开更多
文摘The rRNA genetic locus is found in all prokaryotic organisms, and is highly conservative, although its relatively stable variations are found frequently in different bacteria. The utility of this locus as a taxonomic and phylogenetic tool has been reported widely. This study, aimed at 16S rRNA gene (16S rDNA) and with the help of biomolecular methods, attempted to achieve the goal of rapid identification of common pathogens. In this study, 333 clinical isolated pathogenic bacteria were collected. Two pairs of primers were chosen and labeled with different fluorescent dyes and then used to amplify the genomic DNA extracted from bacteria. The PCR products were then detected by capillary electrophoresis-single strand conformation polymorphism (CE-SSCP). In order to pursue higher resolution and peak-separation effect, a high efficient separating medium, liner polyacrylamidedel (LPA), was put to use in this study. Finally, every bacteria colony generated distinct patterns from each other, which were easily to be used for identification. These results indicated that PCR-CE-SSCP was a rapid identification method for bacterial identification, with the aspects of high efficiency and high precision. Compared with traditional method, this technology is of great utility for clinical use especially for its high sensitivity.
基金Supported by 2010A310011 Henan Provincial Department of Education on Natural Science Research Projects
文摘AIM:To investigate the relationship between c.343A>G and c.2216A>C polymorphism sites in the CDH17 gene and colorectal carcinoma.METHODS:Ninety-three non-consanguineous colorectal carcinoma patients admitted to the Department of Oncology at the First Affiliated Hospital of Zhengzhou University were included in this study.Ninety-three peripheral venous blood samples,of approximately one milliliter from each patient,were collected betweenDecember 2009 and August 2010.The genomic DNA of these peripheral venous blood samples were extracted and purified using a Fermentas Genomic DNA Purification Kit(Fermentas,CA) according to the manufacturer' s protocol.The single nucleotide polymorphisms(SNPs) of the liver-intestine cadherin(CDH17) gene c.343A>G and c.2216A>C were determined by the polymerase chain reaction-single strand conformation polymorphism method(PCR-SSCP) in 93 peripheral venous blood samples from patients suffering with colorectal carcinoma.Typical samples that showed different migration bands in SSCP were confirmed by sequencing.Directed DNA sequencing was used to check the correctness of the genotype results from the PCR-SSCP method.RESULTS:There was a significant association between the c.2216 A>C SNPs of the CDH17 gene and the tumor-node-metastasis(TNM) grade,as well as with lymph node status,in 93 peripheral venous blood samples from colorectal carcinoma patients.The genotype frequencies of A/C,A/A,and C/C were 12.90%,33.33% and 53.76%,respectively.There was a significant correlation between lymph node metastasis,TNM grade,and the genotype distribution(P < 0.05).The C/C genotype raised the risk of lymph node metastasis and the TNM grade.There was a significant difference in the TNM grade and lymph node metastasis between the A/A and C/C genotypes(P = 0.003 and P = 0.013,respectively).Patients with colorectal carcinoma carrying the C allele tended to have a higher risk of lymph node metastasis and have a higher TNM grade.The difference between the TNM grades,as well as the lymph node metastasis of the two alleles,was statistically significant(P < 0.01).CONCLUSION:The SNPs of the CDH17 gene c.2216 A>C might be clinically important in the prognosis of colorectal carcinoma.
文摘By employing different gel components and electrophoresis conditions, the PCR-SSCP(polymerase chain reaction-single-stranded conformation polymorphism) method was used to detect the polymorphism(CCC or CGC ) of codon 72 in exon 4 of p53 gene. The results
文摘Objective: To study the relationship between the polymorphism of drug resistant gene rpoB and drug resistance against rifampicin(RFP) of M. tuberculosis L-forms, and to evaluate its clinical application. Methods: A total of 52 clinical isolated strains of M. tuberculosis L-forms were collected. rpoB gene polymorphism was analyzed by polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) and conventional antimicrobial susceptibility test (AST). Their results were compared. Results: AST results showed that 38 of 52 clinical isolated strains were drug resistance (73.08％),while PCR-SSCP indicated 65.38％ (32/52) rpoB gene polymorphism. There was no statistic significance(χ2= 2.4914) between the 2 methods. Conclusion:Combined the application of PCR-SSCP with AST in detecting rpoB drug resistant gene polymorphism of M. tuberculosis L-form from pneumoconiosis patients with tuberculosis may have advantages at earlier diagnosis and guidance of clinical medications.
文摘Objective To develop a high throughput mutational detection method by mutiple fluorescence-labeled polymerase chain reaction(PCR)products.Methods A total of 27 known mutations including 22 substitutions,3 insertions(1,2 and 7 bp)and 2 deletions(1 and 2 bp)in the hepatocyte nuclear factor(HNF)-4α,glucokinase and HNF-1α genes were tested.During nested PCR,amplified fragments were labeled with three fluorescent dyes.PCR products were visualized with an ABI-377 fluorescence sequencer using 5% glycerol or 10% sucrose in nondenaturing gel conditions.Results Twenty-five of 27 variants(93%)could be detected by combining 5% glycerol and 10% sucrosegel matrix conditions.Twenty-two of 27(82%)and 18 of 27(67%)variants were identified using 5%glycerol and 10% sucrose conditions,respectively.Conclusion This fluorescence-based PCR single strand conformation polymorphism technique represents a simple,non-hazardous,time-saving and sensitive method for high throughput mutation detection.
文摘BACKGROUND: Since single nucleotide polymorphisms (SNPs) can serve as gene markers, polymorphism profiles may help scientists to identify the full collection of genes that contribute to the development of complex diseases such as cancer. The distribution of interleukin-10 (IL-10) promoter polymorphisms in Chinese Han ethnic patients with hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC) was investigated in this study. METHODS: The polymorphisms of IL-10 promoter region were detected by pulymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and sequencing. Sixty-six health controls, 42 patients with HBV infection, 30 HCC patients, and cell line SMMC-7721 were examined this way. RESULTS: Polyrnorphisms of T/C or T/N on-872 site occurred frequently in Han ethnic population. Pulyrnorphisms were detected in HBV and HCC patients and cell line SMMC-7721. The hotspot among the pulymorphisms was inserting base A between-1058 and-1057. CONCLUSION: Polymorphisms of IL-10 promoter in HBV and HCC patients may be associated with HBV infection and HCC development.
基金the National Natural Science Foundation of China, No. 30972500the Natural Science Foundation of Guangdong Province, No. 7301507
文摘Poly （ADP-ribose） polymerase-1 （PARP-1） can exacerbate ischemic brain injury and lessen ischemic neuronal death, which may be associated with PARP-1 polymorphisms. The present study investigated human PARP-1 gene polymorphisms in various Chinese nationalities, the results of which could potentially help in the treatment and prevention of neurologic diseases. Genetic polymorphisms of seven exons in the PARP-1 gene, in 898 Chinese Han, Buyi, Shui, Miao, and Zhuang subjects, were investigated by PCR-single-strand conformation polymorphism. A single-strand conformation polymorphism variant in exons 12, 13, 16, and 17 of the PARP-1 gene was identified in 148 people, with two stationary bands showing three degenerative single strands. Results showed that the PARP-1 gene polymorphisms exist in various nationalities, and may act as a biomarker for susceptibility to disease.
