PCR OPTIMIZATION: IMPROVING OF HUMAN CYTOMEGALOVIRUS (HCMV) PCR
TO ACHIEVE A HIGHLY SENSITIVE DETECTION METHOD†

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AMINI BAVIL OULIAEI S.; SABAHI F.; KARIMI MOHSEN

مرکز اطلاعات علمی Scientific Information Database (SID) - Trusted Source for Research and Academic Resources

Journal Paper

Paper Information

Journal: Iranian Journal of Biotechnology Year:2003 | Volume:1 | Issue:1 Page(s): 59-64

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Information Journal Paper

Title

PCR OPTIMIZATION: IMPROVING OF HUMAN CYTOMEGALOVIRUS (HCMV) PCR TO ACHIEVE A HIGHLY SENSITIVE DETECTION METHOD†

Author(s)

AMINI BAVIL OULIAEI S. | SABAHI F. | KARIMI MOHSEN | Issue Writer Certificate

Keywords

POLYMERASE CHAIN REACTION (PCR)

OPTIMIZATION

HUMAN CYTOMEGALOVIRUS (HCMV)

RENAL TRANSPLANT RECIPIENTS.

Abstract

Polymerase chain reaction (PCR) is a rapid and simple technique with high sensitivity and specificity. In the recent years, PCR has been used for rapid detection of viral nucleic acids, such as Human cytomegalovirus (HCMV), whereas, PCR OPTIMIZATION is an important task to be done, especially before it’s diagnostic application. Annealing temperature, ion concentration (especially Mg2+ ion) and the cycling program and enhancer compounds are important OPTIMIZATION parameters. Peripheral blood leukocytes (PBLs) were isolated from samples collected from renal transplant recipients suffering from severe and symptomatic CMV disease. PBLs DNA was extracted and used for PCR. Annealing temperature and MgCl2 concentration and cycling condition were optimized. Dimethyl sulfoxide (DMSO) and gelatin were checked as enhancer components. The optimized condition obtained through this study was: 1x PCR buffer (20 mM Tris-HCl pH 8.6, 50 mM KCl), 2.5 mM MgCl2, 0.2 mM of each dNTPs, 0.25 µM of each primers, 0.25 unit/25 µl Taq DNA polymerase, 5% DMSO, 500 µg/ml gelatin and 50-150 ng template DNA in 25 µl final volume. PCR was performed as: 95°C 5 min (pre-denaturation), 94°C 50 sec, 58°C 1 min, 72°C 1 min for 35 cycles and 72°C 5 min (final extension). Using these conditions, it was shown that optimized PCR was five fold more sensitive than initial PCR; which can be used for diagnostic application of HCMV in renal transplant patients.

Cites

PRE-SYMPTOMATIC HUMAN CYTOMEGALOVIRUS DISEASE DIAGNOSIS IN RENAL TRANSPLANT RECIPIENTS BY THE VIRUS DNA PCR

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APA: Copy

AMINI BAVIL OULIAEI, S., SABAHI, F., & KARIMI, MOHSEN. (2003). PCR OPTIMIZATION: IMPROVING OF HUMAN CYTOMEGALOVIRUS (HCMV) PCR TO ACHIEVE A HIGHLY SENSITIVE DETECTION METHOD†. IRANIAN JOURNAL OF BIOTECHNOLOGY, 1(1), 59-64. SID. https://sid.ir/paper/527702/en

Vancouver: Copy

AMINI BAVIL OULIAEI S., SABAHI F., KARIMI MOHSEN. PCR OPTIMIZATION: IMPROVING OF HUMAN CYTOMEGALOVIRUS (HCMV) PCR TO ACHIEVE A HIGHLY SENSITIVE DETECTION METHOD†. IRANIAN JOURNAL OF BIOTECHNOLOGY[Internet]. 2003;1(1):59-64. Available from: https://sid.ir/paper/527702/en

IEEE: Copy

S. AMINI BAVIL OULIAEI, F. SABAHI, and MOHSEN KARIMI, “PCR OPTIMIZATION: IMPROVING OF HUMAN CYTOMEGALOVIRUS (HCMV) PCR TO ACHIEVE A HIGHLY SENSITIVE DETECTION METHOD†,” IRANIAN JOURNAL OF BIOTECHNOLOGY, vol. 1, no. 1, pp. 59–64, 2003, [Online]. Available: https://sid.ir/paper/527702/en

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