该研究通过3管型微量MPN计数法(Mini Most Probable Number,mini-MPN)建立了一种快速定量检测食源性沙门氏菌的荧光定量环介导等温扩增(Quantitative Loop-Mediated Isothermal Amplification,qLAMP)方法。依据沙门菌属ttr基因设计了qL...该研究通过3管型微量MPN计数法(Mini Most Probable Number,mini-MPN)建立了一种快速定量检测食源性沙门氏菌的荧光定量环介导等温扩增(Quantitative Loop-Mediated Isothermal Amplification,qLAMP)方法。依据沙门菌属ttr基因设计了qLAMP和荧光定量PCR(Quantitative Polymerase Chain Reaction,qPCR)引物,结合5 h BPW增菌和MPN计数法建立了mini-MPN-qLAMP沙门氏菌快速定量检测方法;使用两种人工污染样品对mini-MPN-qLAMP法进行验证,使用Bland-Altman分析比较不同检测方法检测结果的一致性。结果表明,建立的qLAMP法与qPCR法反应特异性均良好,纯培养时qLAMP法检出限为500 CFU/mL。通过Bland-Altman分析表明所建立的mini-MPN-qLAMP法在速冻乌米饭中检测结果与mini-MPN-qPCR、mini-MPN计数法、平板计数法相比均具有较高的一致性,r^(2)≥0.994,检出限为-0.44 lg MPN/mL;而在速冻鸡胸肉中该法检测结果与mini-MPN-qPCR结果一致性最佳,r^(2)=0.990,检出限为-0.64 lg MPN/mL。肉制品中腐败杂菌会影响mini-MPN计数和平板计数结果,mini-MPN-qLAMP可排除肉制品中腐败杂菌对检测结果的影响。该研究所建立的mini-MPN-qLAMP法简单易行,准确度高,可用于食品中沙门氏菌的快速定量检测。展开更多
Purpose:To analyse the hereditary features of a Chinese pedigree with familial vitreous amyloidosis in Liaoning Province,China,and to investigate the correlation between the clinical appearance of the disease and tran...Purpose:To analyse the hereditary features of a Chinese pedigree with familial vitreous amyloidosis in Liaoning Province,China,and to investigate the correlation between the clinical appearance of the disease and transthyretin(TTR)gene mutation,including the locus and type of TTR gene mutation.Methods:Five patients (10 eyes) from one Chinese family were diagnosed with vitreous amyloidosis between July 1996 and April 2009.Family members were followed up subsequently,and peripheral venous blood was obtained from 13 subjects (including 2 patients,and 11 controls without clinical signs of disease).DNA samples were extracted and 4 exons of the TTR gene were amplified by polymerase chain reaction (PCR).The gene fragments were subjected to sequencing analysis.The results were analyzed with DNAMAN Windows 5.2.2.0 and Chromas sequence chart analysis software,TTR gene exons were compared between affected patients and normal controls.Results:Family pedigree analysis revealed that patients were distributed in three generations.Male and female subjects had equal prevalence,and only one parent of affected patients had signs of disease.TTR gene exon sequencing showed that the sequence of patients was identical to that of normal individuals.No TTR gene mutations were noted in 10 unaffected family members.However,a TTR Gly-54 point mutation in the 2nd exon was detected in two patients and 1 unaffected family member (one of the patients' daughters).Vitreous samples in 4 cases (7 eyes) showed positive Congo red staining,suggesting that these family members suffered from familial vitreous amyloidosis.Conclusion:This pedigree affected with familial vitreous amyloidosis was characterized by autosomal dominant inheritance;.a TTR Gly-54 point mutation in the 2nd exon is presumed to be the cause.This Gly-54 point mutation of the TTR gene is a novel mutation in vitreous amyloidosis.展开更多
文摘该研究通过3管型微量MPN计数法(Mini Most Probable Number,mini-MPN)建立了一种快速定量检测食源性沙门氏菌的荧光定量环介导等温扩增(Quantitative Loop-Mediated Isothermal Amplification,qLAMP)方法。依据沙门菌属ttr基因设计了qLAMP和荧光定量PCR(Quantitative Polymerase Chain Reaction,qPCR)引物,结合5 h BPW增菌和MPN计数法建立了mini-MPN-qLAMP沙门氏菌快速定量检测方法;使用两种人工污染样品对mini-MPN-qLAMP法进行验证,使用Bland-Altman分析比较不同检测方法检测结果的一致性。结果表明,建立的qLAMP法与qPCR法反应特异性均良好,纯培养时qLAMP法检出限为500 CFU/mL。通过Bland-Altman分析表明所建立的mini-MPN-qLAMP法在速冻乌米饭中检测结果与mini-MPN-qPCR、mini-MPN计数法、平板计数法相比均具有较高的一致性,r^(2)≥0.994,检出限为-0.44 lg MPN/mL;而在速冻鸡胸肉中该法检测结果与mini-MPN-qPCR结果一致性最佳,r^(2)=0.990,检出限为-0.64 lg MPN/mL。肉制品中腐败杂菌会影响mini-MPN计数和平板计数结果,mini-MPN-qLAMP可排除肉制品中腐败杂菌对检测结果的影响。该研究所建立的mini-MPN-qLAMP法简单易行,准确度高,可用于食品中沙门氏菌的快速定量检测。
文摘Purpose:To analyse the hereditary features of a Chinese pedigree with familial vitreous amyloidosis in Liaoning Province,China,and to investigate the correlation between the clinical appearance of the disease and transthyretin(TTR)gene mutation,including the locus and type of TTR gene mutation.Methods:Five patients (10 eyes) from one Chinese family were diagnosed with vitreous amyloidosis between July 1996 and April 2009.Family members were followed up subsequently,and peripheral venous blood was obtained from 13 subjects (including 2 patients,and 11 controls without clinical signs of disease).DNA samples were extracted and 4 exons of the TTR gene were amplified by polymerase chain reaction (PCR).The gene fragments were subjected to sequencing analysis.The results were analyzed with DNAMAN Windows 5.2.2.0 and Chromas sequence chart analysis software,TTR gene exons were compared between affected patients and normal controls.Results:Family pedigree analysis revealed that patients were distributed in three generations.Male and female subjects had equal prevalence,and only one parent of affected patients had signs of disease.TTR gene exon sequencing showed that the sequence of patients was identical to that of normal individuals.No TTR gene mutations were noted in 10 unaffected family members.However,a TTR Gly-54 point mutation in the 2nd exon was detected in two patients and 1 unaffected family member (one of the patients' daughters).Vitreous samples in 4 cases (7 eyes) showed positive Congo red staining,suggesting that these family members suffered from familial vitreous amyloidosis.Conclusion:This pedigree affected with familial vitreous amyloidosis was characterized by autosomal dominant inheritance;.a TTR Gly-54 point mutation in the 2nd exon is presumed to be the cause.This Gly-54 point mutation of the TTR gene is a novel mutation in vitreous amyloidosis.