Impact of pesticide treatment on soil microbial population for Different vegetables using ribosomal DNA fingerprinting

Foong, Choon Pin (2007) Impact of pesticide treatment on soil microbial population for Different vegetables using ribosomal DNA fingerprinting. Universiti Malaysia Sabah. (Unpublished)


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Ribosomal DNA fingerprinting using the technique of PCR-DGGE was used in this study to investigate the impact of pesticide treatment on the profile of soil microbial organisms. This technique is based on the separation of polymerase chain reaction-amplified fragments of 16S rDNA, all the same length, by denaturing gradient gel electrophoresis (DGGE). The DNA extraction method used in this study was able to produce large amount of DNA, 1.5 to 3.9 μg of DNA per g of soil and at the same time humic substances were also co-extracted. Dilution of the crude DNA extract to 1000 folds was proved to be able removed inhibitory effects to PCR but high concentration of DNA was a prerequisite. DNA extraction and purification methods were the main factors in determining the subsequent success of PCR, cloning and DGGE. DNA quantification using the agarose gel electrophoresis method was more reliable compared to spectrophotometry method due to the presence of humic substances which would interfere absorbance reading of DNA. In this study, the PCR-cloning strategy was unable to complete due to non-optimizing PCR conditions. Thus, the PCR product concentration was too low and unsuitable to proceed with cloning. Besides, the PCR-DGGE was also unsuccessful to accomplish the aim for DNA fingerprinting construction. Optimization of the conditions of DGGE would be required to obtain a better result.

Item Type: Academic Exercise
Keyword: PCR-DGGE, DNA fingerprinting, pesticide treatment, soil microbial, polymerase chain reaction-amplified
Subjects: Q Science > QH Natural history > QH301-705.5 Biology (General)
Department: SCHOOL > School of Science and Technology
Date Deposited: 29 Apr 2014 13:08
Last Modified: 13 Oct 2017 11:31

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