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Polymerase chain reaction–hybridization method using urease gene sequences for high-throughput Ureaplasma urealyticum and Ureaplasma parvum detection and differentiation.

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Τίτλος: Polymerase chain reaction–hybridization method using urease gene sequences for high-throughput Ureaplasma urealyticum and Ureaplasma parvum detection and differentiation.
Συγγραφείς: Xu, Chen1, Zhang, Nan2, Huo, Qianyu2, Chen, Minghui2, Wang, Rengfeng2, Liu, Zhili2, Li, Xue2, Liu, Yunde2, Bao, Huijing2 kris_10713@126.com
Πηγή: Analytical Biochemistry. Apr2016, Vol. 499, p57-62. 6p.
Θεματικοί όροι: *Polymerase chain reaction, *Urease genetics, *Nucleotide sequence, *DNA probes, *DNA primers
Περίληψη: In this article, we discuss the polymerase chain reaction (PCR)–hybridization assay that we developed for high-throughput simultaneous detection and differentiation of Ureaplasma urealyticum and Ureaplasma parvum using one set of primers and two specific DNA probes based on urease gene nucleotide sequence differences. First, U. urealyticum and U. parvum DNA samples were specifically amplified using one set of biotin-labeled primers. Furthermore, amine-modified DNA probes, which can specifically react with U. urealyticum or U. parvum DNA, were covalently immobilized to a DNA–BIND plate surface. The plate was then incubated with the PCR products to facilitate sequence-specific DNA binding. Horseradish peroxidase–streptavidin conjugation and a colorimetric assay were used. Based on the results, the PCR–hybridization assay we developed can specifically differentiate U. urealyticum and U. parvum with high sensitivity (95%) compared with cultivation (72.5%). Hence, this study demonstrates a new method for high-throughput simultaneous differentiation and detection of U. urealyticum and U. parvum with high sensitivity. Based on these observations, the PCR–hybridization assay developed in this study is ideal for detecting and discriminating U. urealyticum and U. parvum in clinical applications. [ABSTRACT FROM AUTHOR]
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  Label: Title
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  Data: Polymerase chain reaction–hybridization method using urease gene sequences for high-throughput Ureaplasma urealyticum and Ureaplasma parvum detection and differentiation.
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  Data: <searchLink fieldCode="AR" term="%22Xu%2C+Chen%22">Xu, Chen</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Zhang%2C+Nan%22">Zhang, Nan</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Huo%2C+Qianyu%22">Huo, Qianyu</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Chen%2C+Minghui%22">Chen, Minghui</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Wang%2C+Rengfeng%22">Wang, Rengfeng</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Liu%2C+Zhili%22">Liu, Zhili</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Li%2C+Xue%22">Li, Xue</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Liu%2C+Yunde%22">Liu, Yunde</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Bao%2C+Huijing%22">Bao, Huijing</searchLink><relatesTo>2</relatesTo><i> kris_10713@126.com</i>
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  Data: <searchLink fieldCode="JN" term="%22Analytical+Biochemistry%22">Analytical Biochemistry</searchLink>. Apr2016, Vol. 499, p57-62. 6p.
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  Data: *<searchLink fieldCode="DE" term="%22Polymerase+chain+reaction%22">Polymerase chain reaction</searchLink><br />*<searchLink fieldCode="DE" term="%22Urease+genetics%22">Urease genetics</searchLink><br />*<searchLink fieldCode="DE" term="%22Nucleotide+sequence%22">Nucleotide sequence</searchLink><br />*<searchLink fieldCode="DE" term="%22DNA+probes%22">DNA probes</searchLink><br />*<searchLink fieldCode="DE" term="%22DNA+primers%22">DNA primers</searchLink>
– Name: Abstract
  Label: Abstract
  Group: Ab
  Data: In this article, we discuss the polymerase chain reaction (PCR)–hybridization assay that we developed for high-throughput simultaneous detection and differentiation of Ureaplasma urealyticum and Ureaplasma parvum using one set of primers and two specific DNA probes based on urease gene nucleotide sequence differences. First, U. urealyticum and U. parvum DNA samples were specifically amplified using one set of biotin-labeled primers. Furthermore, amine-modified DNA probes, which can specifically react with U. urealyticum or U. parvum DNA, were covalently immobilized to a DNA–BIND plate surface. The plate was then incubated with the PCR products to facilitate sequence-specific DNA binding. Horseradish peroxidase–streptavidin conjugation and a colorimetric assay were used. Based on the results, the PCR–hybridization assay we developed can specifically differentiate U. urealyticum and U. parvum with high sensitivity (95%) compared with cultivation (72.5%). Hence, this study demonstrates a new method for high-throughput simultaneous differentiation and detection of U. urealyticum and U. parvum with high sensitivity. Based on these observations, the PCR–hybridization assay developed in this study is ideal for detecting and discriminating U. urealyticum and U. parvum in clinical applications. [ABSTRACT FROM AUTHOR]
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