Conclusion

Colony PCR or colony multiplex PCR assays proved to be simple to perform as well as being accurate. It is a very sensitive and reproducible method for direct PCR without the need of DNA preparation. Studies based on M. tuberculosis identification using PCR indicated that sonication was the most sensitive method of DNA extraction compared to other protocols such as boiling, nonionic detergents, proteinase K, and freeze-thawing. However, false negative results may occur when viscous specimens are sonicated.[15-18] Afghani and Stutman[9] concluded that boiling is a rapid, sensitive, and specific method of DNA extraction for PCR applications. Unfortunately, accurate estimation of colony PCR sensitivity is difficult. Because the number of bacterial cells used in the PCR to determine the sensitivity of the assay is often adjusted by using the optical density of the suspension, and the actual bacterial cell number per sample may have been higher as a result of the possible clumping of organisms, this will possibly lead to a miscalculated cell count.

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