Isolation and Characterization of Phytase Producing Bacteria from Poultry Farm Soil with Chicken Faeces from Bukit Lagong

Khor, Jing Herng (2018) Isolation and Characterization of Phytase Producing Bacteria from Poultry Farm Soil with Chicken Faeces from Bukit Lagong. Final Year Project (Bachelor), Tunku Abdul Rahman University College.

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Abstract

Phytic acid is one of the most commonly known anti-nutritional factors in the world as it will bind with proteins and minerals to form complexes known as phytate, which is not able to be readily assimilated by the digestive system of many animals due to lack of enzyme phytase. Phytase hydrolyses phytic acid from food and therefore eliminates the anti-nutritional effect. The aim of this research was to isolate phytase-producing bacteria from poultry farm soil and chicken faeces. Phytic acid was extracted from organic wheat bran in this research, and it was shown that wheat bran contained 0.99% (w/w) of phytic acid. Later, bacteria were cultured in phytate specific (PS) agar plates and those successfully produced halo on agar plates were considered as phytase producing bacteria. Of the 21 isolates, five isolates showed halos (clear zones) around the colonies. Two isolates, S1.1 and S1.9 were chosen for phytase kinetic studies. They were both gram positive and rod-shaped bacteria. S1.1 and S1.9 were cultured to produce enzyme phytase. In the condition of 50ºC and at pH5.5, enzyme produced by S1.1 and S1.9 had activity of 8.17 x 10-8 U/ml and 1.32 x 10-7 U/ml respectively. Also, enzyme produced by isolate S1.1 was most active at 40ºC and at pH5.5 while enzyme produced by isolate S1.9 was most active at 30ºC and at pH6.5. Enzyme produced by isolate S1.1 had Vmax value of 0.8571 mM/s and Km value of 136.9932mM while enzyme produced by isolate S1.9 had Vmax value of 0.4614 mM/s and Km value of 56.9649mM. DNA of both isolates was successfully extracted. DNA of isolate S1.9 was amplified using PCR and it had size of about 1500bp. BLAST analysis of 16S rRNA gene sequences of isolate S1.9 revealed 99% sequence similarity to Bacillus sp. strain MTB17 (GenBank Accession No. MH062891.1).