MOLECULAR AND GENETIC CHARACTERIZATION OF SOME Rhizobium leguminosarum bv. vicieae isolates. | ||||
| Journal of Plant Production | ||||
| Article 17, Volume 32, Issue 6, June 2007, Page 4515-4532 PDF (1.12 MB) | ||||
| Document Type: Original Article | ||||
| DOI: 10.21608/jpp.2007.208779 | ||||
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| Authors | ||||
| Howaida M. L. Abd EL- Basit1; Amina A. Hassan2; M. A. H. Youssef2; M. K. Amin2 | ||||
| 1genetic Dept., Fac., Agric., Zagazig Univ. | ||||
| 2Agric. Mictogbiology Dept., Fac. Agric., Zagazig Univ. | ||||
| Abstract | ||||
| Rhizobium leguminosarum bv. vicieae isolates which isolated and fully characterized in Agric. Microbiol. Dept., Agric. Fac., ZagazigUniv. were used in this work to study molecular and genetic diverisity.24 isolates were used to investigate lysogenicity ability. The results showed that 23 isolates from 24 were lysogen with lysogenecity percent 96%. 19 lysogenic isolates were contained more than one different prophage, since the phage released from them was able to lysis the same lysogenic isolate which released from it. The isolates RA21, RF12, RF13 and RR13 were contained one prophage only, since the phage released was not able to lysis the same lysogenic host. Most bacterial isolates were sensitive to streptomycin, ampicillin and chloramphenicol at concentrations from 100 to 2000 μg/ ml.The RR23 isolate was resistance to ampicillin and chloramphenicol at the used concentrations. The RA11 and RR11 isolates were resistanc to ampicillin up to 1500 μg/ ml and chloramphenicol up to 500 μg/ ml. The RK11 and RF12 isolates were resistance to streptomycin up to 500 μg/ ml. Five rhizobiophages were used in this study. These phages were isolated from soil. The host range of these phages was studied by using the bacterial isolates as hosts. The phages A32, R11 and H21 were able to lysis all the used hosts. The phage K23 was lysis 10 from 11 host, while phage F13 was lysis 7 from 10 host. The plaque forming units (pfu/ ml) of these phages were varied, it ranged from 2.2× 106 to 9.67× 1013. The ability of these phages to transduce some antibiotic resistance genes was assessed. The five phages were able to successfully transduce streptomycin and chloramphenicol resistance genes. Transduction frequency ranged from 0.39× 10-8 to 4.5× 10-5 for streptomycin and from 1.25× 10-8 to 1.3× 10-3 for chloramphenicol. Not all phages were able to transduce the ampicillin resistance gene. Also, transducing this marker was not success with all the recipients. Tranceduction frequency ranged from 4.3×10-9 to 9.4 × 10-8. The five phages were able to cotransduce streptomycin and chloramphenicol resistance gene together, tranceduction frequency ranged from 5.0 × 10-5 to 2.97 × 10-3. Random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) analyses were performed by genomic DNA extracted from 10 isolates of Rhizobium leguminosarum bv. Viciae. Five out of 16 arbitrary decamer primers used in this study were informative and detected scoreable polymorphism in banding patterns of RAPD markersbetween these isolates. Each of primers used for analysis of individual isolates amplified different number of bands. Genetic similarity between isolates, calculated as the total number of band differences, were computed. The similarity coefficient value (1.000) was observed among RF31, RF31A, RS3, RZ11, RA21, RH31and RB2 isolates (group A). The similarity coefficient value between RR11 and (group A) was 0.919 and it was 0.973 between (group A) and both of RB2A and RK12 isolates. The similarity coefficient value between RR11 and RB2A was 0.946 while, it was 0.892 between RR11 and RK12.The dendrogram of genetic distances among isolates based on band polymorphisms generated by RAPD-PCR after using the primers clustered the 10 isolates into two main clusters, where RF31, RF31A, RS3, RZ11, RA21, RH31and RB2 isolates (group A) constituted one cluster correlated with RB2A and RK12 while, RR11 isolate formed the second cluster. RR11 isolate was distinguishable by 4 positive unique RAPD markers while, RB2A and RK12 isolates were identified by one positive unique RAPD markers. | ||||
| Keywords | ||||
| Rhizobium; transduction; bacteriophage; RAPD - PCR | ||||
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