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Journal of Zhejiang University SCIENCE A 2000 Vol.1 No.2 P.222-226

http://doi.org/10.1631/jzus.2000.0222


DETECTION OF BACTERIAL DNA BY PCR AND REVERSE HYBRIDIZATION IN THE 16s rRNA GENE


Author(s):  SHANG Shi-qiang, YU Xi-lin, HONG Wen-lan, YU Hui-min, SUN Mei-yue

Affiliation(s):  Dept. of Neonatology, Children's Hospital of Zhejiang University, Hangzhou 310003, China

Corresponding email(s):   YeZH@mail.hz.zj.cn

Key Words:  16s, rDNA, rRNA PCR, bacterial infection, hybridization, neonate


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SHANG Shi-qiang, YU Xi-lin, HONG Wen-lan, YU Hui-min, SUN Mei-yue. DETECTION OF BACTERIAL DNA BY PCR AND REVERSE HYBRIDIZATION IN THE 16s rRNA GENE[J]. Journal of Zhejiang University Science A, 2000, 1(2): 222-226.

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DOI - 10.1631/jzus.2000.0222


Abstract: 
The clinical diagnosis of sepsis is difficult, particularly in neonates. To devise a rapid and reliable method for identifying bacteria in blood and cerebrospinal fluid (CSF), we developed a pair of primers according to the gene encoding 16 s rRNA, found in all bacteria. DNA fragments from different bacterial species and from clinical samples were detected with polymerase chain reaction (PCR), and with reverse hybridization using a universal bacterial probe, a gram-positive probe and a gram-negative probe. Our results showed that a 371 bp DNA fragment was amplified from 20 different bacterial species. No signal was observed when human DNA and viruses were used as templates. The sensitivity could be improved to 10-12 g. All 26 culture-positive clinical samples (22 blood samples and 4 CSF samples), were positive with PCR. The gram-negative and gram-positive probe hybridized to clinical samples and to known bacterial controls, as predicted by Gram

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Reference

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