Asian Journal of Biomedical and Pharmaceutical Sciences

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DNA sequencing by polymer synthesis with variable ratio of deoxynucleotide triphosphate and fluorescent dideoxynucleotide triphosphate

The chemical mechanism of double stranded DNA polymer synthesis is a complex process. The nucleotides are joined together by phosphodiester bond between 5' C of deoxynucleotide triphosphate and O at 3' C of another deoxynucleotide triphosphate with the release of pyrophosphate and hydrogen ion. The synthesis reaction is performed on the template strand based on the complimentary addition of nucleotide attached by hydrogen bonds. The nucleotide adenine A pairs with thymine T and cytosine C pairs with guanine G by double and triple hydrogen bonds respectively. The deoxynucleotide triphosphate dNTP are the synthesizing molecules and fluorescent dideoxynucleotide triphosphate ddNTP are the terminator molecules in the polymer synthesis. The fluorescent ddNTP lacking O at 3' C, terminate the further addition of nucleotide and fluorescence of attached dye is captured by laser light excitation and absorption on the detector. The fluorescence detection read the nucleotide in the synthesis cycle. The DNA sequencing is the reading of nucleotide in the polymer synthesis with the dNTP and fluorescent ddNTP. We have investigated a chemical mechanism of DNA polymer synthesis with variable ratio of dNTP and ddNTP to elucidate the best design of DNA sequencing for robust, error free long read length to scan the genome. The short tandem repeat sequence STR and single nucleotide polymorphism with the reference sequence SNP are molecular markers reside in the size range of 100 to 1000 nucleotides sequence of genome. The variation in the molecular markers is analyzed for human genetic identification and diversity.

Author(s): Negi DS, Shrivastava P, Das SP