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:: Volume 5, Number 4 (Winter 2016) ::
J Fasa Univ Med Sci 2016, 5(4): 542-550 Back to browse issues page
The detection of Klebsiella pneumoniae 16srRNA specific gene by PCR-ELISA technique
Fatemeh Tayebeh1, Jafar Amani 2, Mehdi Moradyar3, Seyed Ali Mirhossaini1
1- Applied Microbiology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
2- Applied Microbiology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran. , jafar.amani@gmail.com
3- Department of Biology, Damghan Azad University, Damghan, Iran.
Abstract:   (4593 Views)

Objective: Klebsiella Pneumoniae is one of the most important infection bacteria of the Enterobacteriaceae family. It is also the second factor of hospital bacteremia due to the gram negative bacteria after Escherichia coli. In this research, this bacteria with the private 16SrDNA gene, was identified through PCR-ELISA technique.

Materials and Methods: In this method the dNTP labeled with Digoxigenin was used for amplifying the specific gene. Streptavidin was loaded in ELISA plate, and then the specific Biotinylated probe was used to connect the Polymerase Chain Reaction (PCR) product. Biotinylated probe was connected to the streptavidin and the amplified gene was attached to the probe. Finally, the digoxigenin antibodies were used to identify the PCR product. The reaction was measured with an ELISA reader.

Result: 16SrDNA of Klebsiella pneumoniae was amplified using the gene specific primers resulted in a fragment of the bp 260. The results of PCR-ELISA showed that this technique does not cross-react with the bacteria in their families. Additionally,  the sensitivity of 6.0 ng was evaluated.

Conclusion: In this research, PCR-ELISA technique was known as an accurate and rapid test for the detection of infection agents by using the specific gene. PCR-ELISA can act as an alternative method instead of time-consuming, less sensitive, and expensive techniques such as culture bacteria in blood agar plates, Macconkey agar, Realtime PCR and differential biochemical tests which are currently used in the research labs.

Keywords: Enterobacteriaceae, Klebsiella pneumoniae, 16SrDNA, PCR-ELISA
Full-Text [PDF 888 kb]   (1257 Downloads)    
Type of Study: Research | Subject: Microbiology
Received: 2015/08/6 | Accepted: 2016/01/4 | Published: 2016/03/7
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Tayebeh F, Amani J, Moradyar M, Mirhossaini S A. The detection of Klebsiella pneumoniae 16srRNA specific gene by PCR-ELISA technique. J Fasa Univ Med Sci. 2016; 5 (4) :542-550
URL: http://journal.fums.ac.ir/article-1-714-en.html
Volume 5, Number 4 (Winter 2016) Back to browse issues page
Journal of Fasa University of Medical Sciences
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