Abstract
Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis(MTB) which mostly affects the lungs. The disease causes deaths of many people every year. There are different methods to detect MTB such as skin test, staining, culture and molecular techniques. Polymerase chain reaction (PCR) is a simple and rapid method for the detection of MTB; however, positive and negative false results reduce the efficiency of this technique. The aim of this study was to design an internal amplification control (IAC) and apply it in MTB PCR test. PCR technique for MTB was optimized by using specific primers for IS6110 gene. The sensitivity and specificity of the test were determined. IAC was constructed with competitive strategy by PCR-cloning technique and the limitation range was determined. The PCR products of MTB and IAC were 245 and 660 bp, respectively on electrophoresis gel. The IC used in PCR testing of MTB is the competitive form in which the range was between 10 million and 10 bacteria and the most suitable internal control concentration for the mix was 1,000 plasmids. After making IC and using it in MTB amplification, it was observed that IC might guarantee the correctness of PCR reaction. Key words: Mycobacterium tuberculosis, polymerase chain reaction (PCR), internal control.
Highlights
Tuberculosis (TB) is a lethal infectious disease which after decades of decline, showed an increase simultaneously with human immunodeficiency virus (HIV) incidence (Cole et al, 1998; Noordhoek et al, 1994)
World Health Organization (WHO) has estimated that if the disease is not controlled, between 2000 and 2020, nearly one billion people worldwide will be infected with this bacterium, 200 million of this number will get sick and 35 million will die (Pauwels et al, 2001; Sohn et al, 2003)
The aim of this study was to design an internal amplification control (IAC) for Mycobacterium tuberculosis (MTB) detection to be used in various samples which may be undetected to cause existence of different inhibitors
Summary
Tuberculosis (TB) is a lethal infectious disease which after decades of decline, showed an increase simultaneously with human immunodeficiency virus (HIV) incidence (Cole et al, 1998; Noordhoek et al, 1994). World Health Organization (WHO) has estimated that if the disease is not controlled, between 2000 and 2020, nearly one billion people worldwide will be infected with this bacterium, 200 million of this number will get sick and 35 million will die (Pauwels et al, 2001; Sohn et al, 2003). For this reason, TB has been announced a global threat by WHO (Fredricks and Relman, 1999). Sputum culture is more sensitive than direct tests; the results
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