دراسة وراثية وبكتريولوجية على بكتريا تثبيت النيتروجين التكافلية Ensifer meliloti == Bacteriological And Genetic Study On Symbiotic Nitrogen Fixing Bacteria Ensifer Meliloti
Author name:
محمد زهير ناجي المرعب
Supervisor name:
حسن فاضل ناجي
General topic:
Biology
Specific topic:
Biotechnologies
Degree:
Master
University:
University of Babylon - College Of Science - Department Of Biology
Language:
Arabic
University location:
Babylon
First pages:
24T2689 - p.pdf
Abstract:
عزلت عشر عزلات من بكتريا Ensifer meliloti من العقد الجذرية لنبات الجت Medicago sativa L. بعد ان جمعت من مناطق مختلفة من محافظة بابل خلال المدة من شهر تشرين الثاني/2012 لغاية شهر شباط/2013. شخصت العزلات جزيئيا بتقنية تفاعل البلمرة السلسلي فضلا عن طرق ال | Ten isolates of Ensifer meliloti were isolated from root nodules of Medicago sativa L. collected from different areas of Babylon province during a period from November - 2012 to February - 2013. The isolates were molecular identified using polymerase chain reaction technique, in addition to traditional methods (morphological and biochemical). All isolates were harboured nodbox and mucR genes and produced cyclic ? - (1, 2) glucans, cellulose fibers and lipopolysaccharide.The ability of isolates to grow at different degrees of pH, temperatures and different concentrations of sodium chloride were tested. It was found that the optimum pH for growing of Em1, Em3, Em4, Em6 and Em8 isolates was pH7, while for the isolates Em2, Em5, Em7, Em9 and Em10 was pH8. The optimum temperature for growing of Em1, Em3, Em5, Em7, Em8 and Em9 isolates was 30?C, while the optimal growth of Em2, Em 4 and Em6 isolates was 35?C, Em10 isolate showed significantly extremism to grow at temperture reached to 45?C. The optimal percentages of NaCl for growing of Em1, Em3, Em4, Em5, Em6, Em8 and Em9 isolates were 2%, for Em7 isolate was 3% and for Em2 and Em10 isolates were 4%. The effect of different degrees of pH on symbiotic properties was studied. Em1 and Em8 isolates that grew at pH6.5 ; all isolates that grew at pH 7, pH 7.5 and pH 8 were able to infect the root hairs of alfalfa plants. The isolates that grew at pH6.5 (except Em1 and Em8 isolates) and all isolates that grew at pH8.5 were failed to infect the root hairs and did not formed nodules and resembled to the control plants (without inoculum).Different concentrations of NaCl on symbiosis was studied, it was found that all isolates that grew at 2% and Em2 and Em10 isolates that grew at 3% formed nodule, while Em1, Em3, Em4, Em5, Em6, Em7, Em8 and Em9 isolates that grew at 3% and all isolates that grew at 4%, 5% and 6% did not elicited nodules on the roots of alfalfa plant. Light microscopic observations of root hairs inoculated with Ensifer meliloti isolates showed the root hair deformations like curling, waviness, swelling and formation of shepherd`s crook and infection thread. These consequent changes lead to emerge the mature nodules having five distinct zones viz., distal meristematic, infection, inter, nitrogen - fixing and proximal senescence zones.The gel electrophoresis results of plasmid DNA of isolates showed that these isolates harboured two megaplasmids. When the isolates exposed to extreme factors like pH, temperture and NaCl, it was found that the isolates lost these plasmids and its ability to elicite the nodules and failed to fix the nitrogen. This study showed that the nitrogen fixing bacteria had essential role to support the plant growth and increase their productivity, and recommended to use the isolate Em10 as example for biofertilizers and the best alternative to the chemical fertilizers
