PCR Analysis: Detection Of Bacteriophage Contamination In Biotechnology Used To Produce Recombinant drugs

sambasivarao yaragalla

Abstract


Contamination is a major drawback of the world in biotechnology field and it has cost of losing important biological products like recombinant drugs or valuable research. The causative agents are different chemicals, invertebrates, bacteriophages, bacteria, fungi, parasites, viral species and even other cell lines. In this study, E. coli  BL-21 DE3 (DE3 has λ - bacteriophage genome) which is cloned with therapeutic drug were studied during 2 years (2008-2010) to detect their contaminations and the causative organisms. Samples were examined for bacteriophage contamination using conventional molecular biology techniques. Using polymerase chain reaction (PCR) technology, primers were specifically designed for lambda phage DE3 genome in E. coli  BL-21 (DE3)  and bacteriophage was detected in positive control. No bacteriophage was detected in any of the negative control samples. PCR results were confirmed by virus isolation experiments performed with PCR-positive and negative samples. This indicates λDE3 from E. coli  BL-21 (DE3)   is expressed and this led to cell lysis. The results obtained in this study furnish evidence suggesting that the employed assay system constitutes an effective tool for the detection of contamination in culture systems. To sum up, PCR represents a sensitive, specific, accurate, inexpensive, and quick bacteriophage detection assay that is suitable for the routine screening of cultures.


Keywords


E. coli BL-21 (DE3); PCR; Bacteriophage

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