BACKGROUND Quantitative fluorescent polymerase chain reaction(QF-PCR)is a rapid prenatal diagnostic method for abnormalities on chromosomes 21,18,and 13 and sex chromosomal aneuploidy.However,the value of QF-PCR in di...BACKGROUND Quantitative fluorescent polymerase chain reaction(QF-PCR)is a rapid prenatal diagnostic method for abnormalities on chromosomes 21,18,and 13 and sex chromosomal aneuploidy.However,the value of QF-PCR in diagnosing chromosomal structural abnormalities is limited.In this article,we report a confusing QF-PCR finding in a pregnant woman who underwent amniocentesis.CASE SUMMARY The short tandem repeat marker AMXY(Xp22.2/Yp11.2)located on the sex chromosome exhibited a trisomic biallelic pattern,indicating that the karyotype of the fetus might be 47,XYY.Chromosome analysis performed on cultured amniocytes showed a normal male karyotype of the fetus.Copy number variation sequencing confirmed a 500 kb duplication at Yp11.2-Yp11.2(chrY:6610001_7110000)and a 250 kb duplication at Yp11.2-Yp11.2(chrY:7110001_7360000).CONCLUSION In conclusion,the comprehensive application of different methods could achieve a higher detection rate and accuracy for the prenatal diagnosis of chromosomal disorders through chromosomal testing.展开更多
AIM: To identify and understand the regular distribution pattern and primary penetration site for Salmonella enteritidis (S. enteritidis) in the gastrointestinal tract. METHODS: Based on the species-specific DNA seque...AIM: To identify and understand the regular distribution pattern and primary penetration site for Salmonella enteritidis (S. enteritidis) in the gastrointestinal tract. METHODS: Based on the species-specific DNA sequence of S. enteritidis from GenBank, a species-specific real- time, fluorescence-based quantitative polymerase chain reaction (FQ-PCR) was developed for the detection of S. enteritidis. We used this assay to detect genomic DNA of S. enteritidis in the gastrointestinal tract, including duodenum, jejunum, ileum, cecum, colon, rectum, esophagus and stomach, from mice after oral infection. RESULTS: S. enteritidis was consistently detected in all segments of the gastrointestinal tract. The jejunum and ileum were positive at 8 h post inoculation, and the final organ to show a positive result was the stomach at 18 h post inoculation. The copy number of S. enteritidis DNA in each tissue reached a peak at 24-36 h post inoculation, with the jejunum, ileum and cecum containing high concentrations of S. enteritidis, whereas the duodenum, colon, rectum, stomach and esophagus had low concentrations. S. enteritidis began to decrease and vanished at 2 d post inoculation, but it was still present up to 5 d post inoculation in the jejunum, ileum andcecum, without causing apparent symptoms. By 5 d post inoculation, the cecum had significantly higher numbers of S. enteritidis than any of the other areas (P < 0.01), and this appeared to reflect its function as a repository for S. enteritidis. CONCLUSION: The results provided significant data for clarifying the pathogenic mechanism of S. enteritidis in the gastrointestinal tract, and showed that the jejunum, ileum and cecum are the primary sites of invasion in normal mice after oral infection. This study will help to further understanding of the mechanisms of action of S. enteritidis.展开更多
AIM： To identify and understand the regular distribution pattern for Salmonella enteritidis （S. enteritidis） in the internal organs of mice after an oral challenge over a 3 wk period. METHODS： Assays based on the ...AIM： To identify and understand the regular distribution pattern for Salmonella enteritidis （S. enteritidis） in the internal organs of mice after an oral challenge over a 3 wk period. METHODS： Assays based on the serovar-specific DNA sequence of S. enteritidis from GenBank, and a serovar-specific real-time, fluorescence-based quantitative polymerase chain reaction （FQ-PCR） were developed for the detection of S. enteritidis. We used this assay to detect genomic DNA of S. enteritidis in the blood and the internal organs, including heart, liver, spleen, kidney, pancreas, and gallbladder, from mice after oral challenge at different time points respectively.RESULTS： The results showed that the spleen was positive at 12 h post inoculation （PI）, and the blood was at 14 h PI. The organism was detected in the liver and heart at 16 h PI, the pancreas was positive at 20 h PI, and the final organs to show positive results were the kidney and gallbladder at 22 h PI. The copy number of S. enteritidis DNA in each tissue reached a peak at 24-36 h PI, with the liver and spleen containing high concentrations of S. enteritidis, whereas the blood, heart, kidney, pancreas, and gallbladder had low concentrations. S. enteritidis populations began to decrease and were not detectable at 3 d PI, but were still present up to 12 d PI in the gallbladder, 2 wk for the liver, and 3 wk for the spleen without causing apparent symptoms.CONCLUSION： The results provided significant data for understanding the life cycle of S. enteritidis in the internal organs, and showed that the liver and spleen may be the primary sites for setting itself up as a commensa over a long time after oral challenge. Interestingly, it may be the first time reported that the gallbladder is a site of carriage for S. enteritidis over a 12 d period. This study will help to understand the mechanisms of action of S. enteriCdis infection in vivo.展开更多
INTRODUCTIONAcute narcotizing pancreatitis usually takes a severe clinical course and is associated with multiple organ dysfunction .With the further understanding of pathophysiological events of acute pancreatisis an...INTRODUCTIONAcute narcotizing pancreatitis usually takes a severe clinical course and is associated with multiple organ dysfunction .With the further understanding of pathophysiological events of acute pancreatisis and the therapeutic measuses taken by the clinicians ,the patients can pass through the critical carry stages ,and then the septic complication caused by rtanslocated bacteria, mostly gram-negative microbes from the intestines ensues[1].展开更多
A quantitative real time reverse-transcription polymerase chain reaction （qRT-PCR） assay with specific primers recommended by the World Health Organization （WHO） has been widely used successfully for detection and...A quantitative real time reverse-transcription polymerase chain reaction （qRT-PCR） assay with specific primers recommended by the World Health Organization （WHO） has been widely used successfully for detection and monitoring of the pandemic H1N1/2009 influenza A virus. In this study, we report the design and characterization of a novel set of primers to be used in a qRT-PCR assay for detecting the pandemic H1N1/2009 virus. The newly designed primers target three regions that are highly conserved among the hemagglutinin （HA） genes of the pandemic HlN1/2009 viruses and are different from those targeted by the WHO-recommended primers. The qRT-PCR assays with the newly designed primers are highly specific, and as specific as the WHO-recommended primers for detecting pandemic H1N1/2009 viruses and other influenza viruses including influenza B viruses and influenza A viruses of human, swine, and raccoon dog origin. Furthermore, the qRT-PCR assays with the newly designed primers appeared to be at least 10-fold more sensitive than those with the WHO-recommended primers as the detection limits of the assays with our primers and the WHO-recommended primers were 2.5 and 25 copies of target RNA per reaction, respectively. When tested with 83 clinical samples, 32 were detected to be positive using the qRT-PCR assays with our designed primers, while only 25 were positive by the assays with the WHO-recommended primers. These results suggest that the qRT-PCR system with the newly designed primers represent a highly sensitive assay for diagnosis of the pandemic HIN1/2009 virus infection.展开更多
Aureococcus anophagefferens, a small pelagophyte algae, has caused brown tide blooms in coastal waters of Qinhuangdao in recent years, presenting significant negative impacts on the shellfish mariculture industry. Und...Aureococcus anophagefferens, a small pelagophyte algae, has caused brown tide blooms in coastal waters of Qinhuangdao in recent years, presenting significant negative impacts on the shellfish mariculture industry. Under standard light microscopy, it is visually indistinguishable from other small algae in field samples due to its extremely small size. In this study, quantitative polymerase chain reaction（q PCR） based on 18 S r DNA sequences was developed and used to detect and enumerate A. anophagefferens. A linear regression（R2 = 0.91） was generated based on cycle thresholds value（Ct） versus known concentrations of A. anophagefferens. Twenty-two field samples collected in coastal waters of Qinhuangdao were subjected to DNA extraction and then analyzed using q PCR. Results showed that A. anophagefferens had a wide distribution in coastal waters along Qinhuangdao. Elevated A. anophagefferens abundance, category 3 brown tide blooms（〉200 000 cells/m L） occurred at Dongshan Beach and Tiger-stone Beach in August in 2013. In shellfish mariculture areas along coastal waters of Qinhuangdao, 4 stations had category 3 blooms, and 6 stations had category 2 blooms（35 000–200 000 cells/m L） in August and all stations had category 1 blooms（〉0 to ≤35 000 cells/m L） in October. Quantitative PCR allows for detection of A. anophagefferens cells at low levels in filed samples, which is essential to effective management and prediction of brown tide blooms.展开更多
Objective:The aim of this study was to detect the expression level of Metadherin (MTDH) in peripheral blood of the breast cancer patients by real-time fluorescence quantitative polymerase chain reaction (PCR),and to e...Objective:The aim of this study was to detect the expression level of Metadherin (MTDH) in peripheral blood of the breast cancer patients by real-time fluorescence quantitative polymerase chain reaction (PCR),and to explore the relationship between expression of Metadherin gene in the patients peripheral blood and the clinic-pathological features in breast cancer. Methods:Real-time fluorescence quantitative polymerase chain reaction was employed to determine the expression level of Metadherin gene in 80 peripheral blood samples of breast cancer patients and healthy donors. Results:The expression of Metadherin gene in breast cancer patients peripheral blood were positive,in which 34 breast cancer patients were highly expressed,accounting for 55.7%,while the expression of Metadherin gene in normal females peripheral blood were negative,there was statistical significance (Ratio = 2.02±0.81,P < 0.05); Ratio of the Metadherin expression in breast cancer patients peripheral blood and the glyceraldehyde-3-phosphate dehydrogenase expression was 1.15 ± 0.36. REST software analysis showed that the expression of Metadherin gene was significantly up-regulated in breast cancer. Conclusion:The SYBR Green I quantitative real-time polymerase chain reaction method can successfully detect the expression level of Metadherin gene. Expression level of Metadherin gene in breast cancer patients peripheral blood is closely related to survival,and it maybe involved in the development of breast cancer and used as an indicator of prognosis.展开更多
文摘BACKGROUND Quantitative fluorescent polymerase chain reaction(QF-PCR)is a rapid prenatal diagnostic method for abnormalities on chromosomes 21,18,and 13 and sex chromosomal aneuploidy.However,the value of QF-PCR in diagnosing chromosomal structural abnormalities is limited.In this article,we report a confusing QF-PCR finding in a pregnant woman who underwent amniocentesis.CASE SUMMARY The short tandem repeat marker AMXY(Xp22.2/Yp11.2)located on the sex chromosome exhibited a trisomic biallelic pattern,indicating that the karyotype of the fetus might be 47,XYY.Chromosome analysis performed on cultured amniocytes showed a normal male karyotype of the fetus.Copy number variation sequencing confirmed a 500 kb duplication at Yp11.2-Yp11.2(chrY:6610001_7110000)and a 250 kb duplication at Yp11.2-Yp11.2(chrY:7110001_7360000).CONCLUSION In conclusion,the comprehensive application of different methods could achieve a higher detection rate and accuracy for the prenatal diagnosis of chromosomal disorders through chromosomal testing.
基金Supported by The National Key Technology R&D Program of China, No. 2004B A901A03Program for Chang Jiang Scholars and Innovative Research Team in University, No. IRTO753+2 种基金Program for New Century Excellent Talents in University, No. NCET-04-0906Sichuan Province Basic Research Program, No. 04JY0290061Program for Key Disciplines Construction of Sichuan Province, No. SZD0418
文摘AIM: To identify and understand the regular distribution pattern and primary penetration site for Salmonella enteritidis (S. enteritidis) in the gastrointestinal tract. METHODS: Based on the species-specific DNA sequence of S. enteritidis from GenBank, a species-specific real- time, fluorescence-based quantitative polymerase chain reaction (FQ-PCR) was developed for the detection of S. enteritidis. We used this assay to detect genomic DNA of S. enteritidis in the gastrointestinal tract, including duodenum, jejunum, ileum, cecum, colon, rectum, esophagus and stomach, from mice after oral infection. RESULTS: S. enteritidis was consistently detected in all segments of the gastrointestinal tract. The jejunum and ileum were positive at 8 h post inoculation, and the final organ to show a positive result was the stomach at 18 h post inoculation. The copy number of S. enteritidis DNA in each tissue reached a peak at 24-36 h post inoculation, with the jejunum, ileum and cecum containing high concentrations of S. enteritidis, whereas the duodenum, colon, rectum, stomach and esophagus had low concentrations. S. enteritidis began to decrease and vanished at 2 d post inoculation, but it was still present up to 5 d post inoculation in the jejunum, ileum andcecum, without causing apparent symptoms. By 5 d post inoculation, the cecum had significantly higher numbers of S. enteritidis than any of the other areas (P < 0.01), and this appeared to reflect its function as a repository for S. enteritidis. CONCLUSION: The results provided significant data for clarifying the pathogenic mechanism of S. enteritidis in the gastrointestinal tract, and showed that the jejunum, ileum and cecum are the primary sites of invasion in normal mice after oral infection. This study will help to further understanding of the mechanisms of action of S. enteritidis.
基金The National Key Technology R&D Program of China, No. 2004BA901A03National Scientific and Technical Support Program, No. 2007Z06-017+3 种基金The Cultivation Fund of the Key Scientific and Technical Innovation Project & Ministry of Education of China, No. 706050Program for New Century Excellent Talents in University, No. NCET-04-0906/NCET-06-0818Sichuan Province Basic Research Program, No. 04JY0290061/07JY029-017Program for Key DisciplinesConstruction of Sichuan Province No. SZD0418
文摘AIM： To identify and understand the regular distribution pattern for Salmonella enteritidis （S. enteritidis） in the internal organs of mice after an oral challenge over a 3 wk period. METHODS： Assays based on the serovar-specific DNA sequence of S. enteritidis from GenBank, and a serovar-specific real-time, fluorescence-based quantitative polymerase chain reaction （FQ-PCR） were developed for the detection of S. enteritidis. We used this assay to detect genomic DNA of S. enteritidis in the blood and the internal organs, including heart, liver, spleen, kidney, pancreas, and gallbladder, from mice after oral challenge at different time points respectively.RESULTS： The results showed that the spleen was positive at 12 h post inoculation （PI）, and the blood was at 14 h PI. The organism was detected in the liver and heart at 16 h PI, the pancreas was positive at 20 h PI, and the final organs to show positive results were the kidney and gallbladder at 22 h PI. The copy number of S. enteritidis DNA in each tissue reached a peak at 24-36 h PI, with the liver and spleen containing high concentrations of S. enteritidis, whereas the blood, heart, kidney, pancreas, and gallbladder had low concentrations. S. enteritidis populations began to decrease and were not detectable at 3 d PI, but were still present up to 12 d PI in the gallbladder, 2 wk for the liver, and 3 wk for the spleen without causing apparent symptoms.CONCLUSION： The results provided significant data for understanding the life cycle of S. enteritidis in the internal organs, and showed that the liver and spleen may be the primary sites for setting itself up as a commensa over a long time after oral challenge. Interestingly, it may be the first time reported that the gallbladder is a site of carriage for S. enteritidis over a 12 d period. This study will help to understand the mechanisms of action of S. enteriCdis infection in vivo.
文摘INTRODUCTIONAcute narcotizing pancreatitis usually takes a severe clinical course and is associated with multiple organ dysfunction .With the further understanding of pathophysiological events of acute pancreatisis and the therapeutic measuses taken by the clinicians ,the patients can pass through the critical carry stages ,and then the septic complication caused by rtanslocated bacteria, mostly gram-negative microbes from the intestines ensues[1].
基金supported by grants from National Basic Research Program of China (No.2011CB504800)National Natural Science Foundation of China (No. 31100128 and 81030031)+3 种基金National Mega Project on Major Drug Development (2009ZX09103-678)National Small Business Innovation and Research (SBIR) Program of Chinathe Technology R & D Program of Jiangsu Province, China (BG20077035 and BG2008662)NIH (RO1-AI041927,RO1-AI050468, RO1-DE014145, and RO1-DE014842)
文摘A quantitative real time reverse-transcription polymerase chain reaction （qRT-PCR） assay with specific primers recommended by the World Health Organization （WHO） has been widely used successfully for detection and monitoring of the pandemic H1N1/2009 influenza A virus. In this study, we report the design and characterization of a novel set of primers to be used in a qRT-PCR assay for detecting the pandemic H1N1/2009 virus. The newly designed primers target three regions that are highly conserved among the hemagglutinin （HA） genes of the pandemic HlN1/2009 viruses and are different from those targeted by the WHO-recommended primers. The qRT-PCR assays with the newly designed primers are highly specific, and as specific as the WHO-recommended primers for detecting pandemic H1N1/2009 viruses and other influenza viruses including influenza B viruses and influenza A viruses of human, swine, and raccoon dog origin. Furthermore, the qRT-PCR assays with the newly designed primers appeared to be at least 10-fold more sensitive than those with the WHO-recommended primers as the detection limits of the assays with our primers and the WHO-recommended primers were 2.5 and 25 copies of target RNA per reaction, respectively. When tested with 83 clinical samples, 32 were detected to be positive using the qRT-PCR assays with our designed primers, while only 25 were positive by the assays with the WHO-recommended primers. These results suggest that the qRT-PCR system with the newly designed primers represent a highly sensitive assay for diagnosis of the pandemic HIN1/2009 virus infection.
基金The National Natural Science Foundation of China under contract No.40906081the Public Science and Technology Research Funds Projects of Ocean under contract Nos 2011418021 and 201305030Integrated Environmental Governances and Restoration in Beidaihe Coast of Hebei Province
文摘Aureococcus anophagefferens, a small pelagophyte algae, has caused brown tide blooms in coastal waters of Qinhuangdao in recent years, presenting significant negative impacts on the shellfish mariculture industry. Under standard light microscopy, it is visually indistinguishable from other small algae in field samples due to its extremely small size. In this study, quantitative polymerase chain reaction（q PCR） based on 18 S r DNA sequences was developed and used to detect and enumerate A. anophagefferens. A linear regression（R2 = 0.91） was generated based on cycle thresholds value（Ct） versus known concentrations of A. anophagefferens. Twenty-two field samples collected in coastal waters of Qinhuangdao were subjected to DNA extraction and then analyzed using q PCR. Results showed that A. anophagefferens had a wide distribution in coastal waters along Qinhuangdao. Elevated A. anophagefferens abundance, category 3 brown tide blooms（〉200 000 cells/m L） occurred at Dongshan Beach and Tiger-stone Beach in August in 2013. In shellfish mariculture areas along coastal waters of Qinhuangdao, 4 stations had category 3 blooms, and 6 stations had category 2 blooms（35 000–200 000 cells/m L） in August and all stations had category 1 blooms（〉0 to ≤35 000 cells/m L） in October. Quantitative PCR allows for detection of A. anophagefferens cells at low levels in filed samples, which is essential to effective management and prediction of brown tide blooms.
文摘Objective:The aim of this study was to detect the expression level of Metadherin (MTDH) in peripheral blood of the breast cancer patients by real-time fluorescence quantitative polymerase chain reaction (PCR),and to explore the relationship between expression of Metadherin gene in the patients peripheral blood and the clinic-pathological features in breast cancer. Methods:Real-time fluorescence quantitative polymerase chain reaction was employed to determine the expression level of Metadherin gene in 80 peripheral blood samples of breast cancer patients and healthy donors. Results:The expression of Metadherin gene in breast cancer patients peripheral blood were positive,in which 34 breast cancer patients were highly expressed,accounting for 55.7%,while the expression of Metadherin gene in normal females peripheral blood were negative,there was statistical significance (Ratio = 2.02±0.81,P < 0.05); Ratio of the Metadherin expression in breast cancer patients peripheral blood and the glyceraldehyde-3-phosphate dehydrogenase expression was 1.15 ± 0.36. REST software analysis showed that the expression of Metadherin gene was significantly up-regulated in breast cancer. Conclusion:The SYBR Green I quantitative real-time polymerase chain reaction method can successfully detect the expression level of Metadherin gene. Expression level of Metadherin gene in breast cancer patients peripheral blood is closely related to survival,and it maybe involved in the development of breast cancer and used as an indicator of prognosis.
