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Jawaharlal Nehru Krishi Vishwa Vidyalaya, Jabalpur

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20131
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Subject: Molecular Biology and Biotechnology × Author: Shinde, Vishwajeet × End date: 2013 × Start date: 2013 × Thesis Type: M.Sc ×
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    ThesisItemOpen Access
    Amplification of Phy Gene From Bacillus Species sp. Isolated from Soil
    (JNKVV, 2013) Shinde, Vishwajeet; Tantwai, Keerti
    ABSTRACT Phytases (myo-inositol hexakisphosphate phosphohydrolase), also known as phytase-degrading enzymes, hydrolyses phytate to myo-inositol and phosphoric acid in stepwise manner forming myo-inositol phosphate intermediates. Phytases are widespread in nature because they can be found in animals, plants and microorganisms. Phytases produced biotechnologically have emerged as indispensable entities in feed and food industry for increased bioavailability of essential minerals. In plant-derived foods, phytate acts as an antinutritive factor by causing mineral deficiency due to efficient chelation of metal ions and by forming complexes with proteins. Unlike ruminates, monogastric animals such as pigs and poultry lack phytase-producing microorganisms in their gut. The addition of microbial phytase to the feedstuff for monogastric animals can substantially improve phosphorus utilization, reduce excretion of phosphorus in the faces and counteract the antinutritional properties of phytase. Ongoing interest in isolation of new bacterial strains producing phytase and optimization of this enzyme produces the recombinant feed poses the need for further investigation. The present investigation, entitled “Amplification of phy gene from Bacillus spp. isolated from soil” deals with the amplification and sequencing of phy gene with the help of phy gene specific primers. Genomic DNA was isolated from the single colonies of all the four bacterial isolates (BS 1, BS 2, BS 3 and BS 4). Then the purified DNA was used for the PCR amplification at 58ºC annealing temperature by using phy gene specific primers. The amplified PCR product showed fragment of ~650 bp in size. The amplified sequences of phy gene were sequenced and sequencing revealed the length of 634 and 635 bp from BS2 and BS4 isolates respectively. In silico analysis was carried out by using different software’s which showed 69.7% homology with B. subtilis strains. Phylogenetic tree also justified similarities with other Bacillus spp. Analysis of ORF and amino acid sequence was carried out, the nucleotide sequences showed the complete open reading frame and deduced amino acid sequences showed two sites of stop codon.