Asian Journal of Microbiology, Biotechnology & Environmental Sciences Paper

Vol 20, Issue 2, 2018; Page No.(643-654)

GENETIC COMPARISON AMONG HIGH PHOSPHATESOLUBILIZING MUTANTS OF ENTEROBACTER CLOCAE VIA RAPD-PCR ANALYSIS

EL-HAMSHARY, O. I. M., EFFAT A. M. SOLIMAN, HIBA J.K. AL- MUSALAM AND ALAWIAH ALHEBSHI

Abstract

Phosphate-solubilizing bacterial isolate was strains from the rhizospheric of Purslane (Portulaca oleracea) plant cultivated in Al-Taief, Saudi Arabia. It was isolated based on the formation of clear zone around its colony on Pikoviskya medium. The isolate was identified using biochemical profiling as E. cloacae A36 strain and screened on the basis of solubilization of inorganic tricalcium phosphate using Pikoviskya and National Botanical Research Institute’s phosphate media. The isolate was subjected to two different mutagens, Ethidium bromide (EtBr) and UV irradiation to enhance the phosphate solubilizing activity. In case of EtBr treatment, the survival percentage was decreased with the increase of exposure time and became 0.43 % at exposure time of 120 min. No mutants were obtained when colonies were tested and compared with wild type strain. Results revealed that UV mutagenesis was found more effective as compared to EtBr and the survival rate were decreased gradually whatever with the increase of exposure time and radiation intensities, i.e., whether distances were 5 or 11.5 cm. Results showed that the decreasing in survival percentages was highest in a distance of 5 cm when compared to 11.5 cm and survival was sharply reduced and became 0% at exposure time 10 min. in both distances. It also became 0% at exposure time 6 and 8 min. at a distance of 5 cm. All tested colonies obtained after UV irradiation treatment at a distance of 11.5 cm did not exhibit high efficiency in phosphate solubilization compared with distance of 5 cm. A total of one hundred colonies obtained after 3 min. of UV irradiation treatment at 5 cm distance and were assayed for phosphate solubilizing. Twenty-seven mutants exhibited higher efficiencies. Eight mutants (M36, M35, M40, M29, M27, 8M4, M14 and 2M4) were selected based on the most efficient phosphate solubilization at 300C on Pikovskaya’s and National Botanical Research Institute’s phosphate agar media. Results revealed that all selected mutants were able to solubilize TCP on solid culture state by forming clear zone, with different degree of solubilization. RAPD-PCR protocol on ten mutants in comparison with the wild type using random primers lead to detect of some changes in RAPD profiles which confirm the genetic variability’s of E. cloacae mutants.

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