Actinomycete Strains

STUDIES ON THE DISTRIBUTION OF PROTEINS DONE IN MARINE ACTINOMYCETE STRAINS

Sangeetha.V.S1, Shabi Ruskin R1, Praseetha.P.K2 Chandraja.C.V3,Anupa.M.P.1

1Department of Biotechnology,

2Department of Bioinformatics

3Department of Microbiology

Noorul Islam College of Arts and Science, Kumaracoil

ABSTRACT

Various Actinomycete strains were observed and 5 different strains with distinct morphological characters were selected for the study. The protein concentration was determined for each strain at different temperatures (32°C, 37°C and 40°C) and at different pH (6.5, 6.8, 7, 7.2 and 7.4). The protein concentration was also determined at different growth periods. The 92 hrs cultures of Actinomycete strain NIC5 showed least protein concentration

Then the seaweeds were macerated well with distilled water by using a mortar and pestle. The sample was serially diluted and spread plate technique was performed. After 7 days of incubation at 28°C, the total number of bacterial, fungal and Actinomycetes colonies in each petridish was counted and the randomly selected Actinomycetes colonies were sub cultured in potato dextrose agar slants. After obtaining satisfactory growth, the slants were kept in a refrigerator (4°C) till further analysis was carried out.

4. Pure Culture and Stock Culture of Actinomycetes The Potato Dextrose Agar slants were prepared and colonies were isolated from the plates and were inoculated into the Potato Dextrose Agar slant. The slants were incubated at room temperature for 5-7 days. After incubation, the colonies were observed and the plates were stored in the refrigerator.

5. Mass Culture The colonies were isolated from the potato Dextrose Agar slant and inoculated into the 100ml sterile Potato Dextrose Agar broth. Then incubated at room temperature for 4-5 days. After incubation the cells were cultured in Potato Dextrose Agar

Estimation of protein by Lowry’s method The total proteins present in the intracellular and extra cellular fluids were determined by Lowry’s method. The OD values were plotted in along with the standard values on to graph sheet to obtain the concentration of protein in the samples.

8. Assay of Proteases

The protease activity of all selected Actinomycete strains were estimated at 36 & 48 hours of growth. Into a series of ten test tubes 1ml of casein was taken and another control tube containing 2ml of casein was taken. Then 6.2ml of enzyme was added in all the tubes. Then it was incubated at 370C for 10 minutes. After incubation 1ml of 12% TCA was added, and the mixture was cooled rapidly in ice, and centrifuged at 4000 rpm for 10minutes of the supernatant was collected. After this 5 ml of Reagent c and 1.5ml of Folin’s reagent was added to all the tubes. Then it was incubated at room temperature for 10 minutes. The blue colour developed and it was read at 650nm. The OD values were plotted in along with the standard values onto a graph sheet to obtain the enzyme activity of protease in the