5242
RASHA GOMMA SAID SALIM
GENETIC CONSTRUCTION OF NEW STRAINS OF LACTIC ACID BACTERIA
Bacteriocin, Lactic acid bacteria, 16S rRNA gene, RAPD PCR Conjugation, Bacteriocin genes , Cloning.
Ten local isolates of lactic acid bacteria (LAB) able to produce bacteriocin were purified from traditional dairy products and then identified morphologically. These isolates were tested for activity of bacteriocin production against a wide range of food spoilage pathogens and termed as I1 to I10. Three different isolates I1, I2 and I3 were selected as the best bacterial isolates that gave a high activity of the bacteriocin production. The morphological, biochemical and molecular identifications based on (16s rRNA gene). It characterized I1 as E. faecium, I2 as E. faecium and I3 as Pediococcus pentosaceus. The sequences of these isolates were deposited in GenBank database under accession numbers LC063691, LC063692 and LC063861. The genetic improvement was done via conjugation as one important method to obtain strains have high bacteriocin production and activity compared with parents. Genetic improvement of selected bacterial isolates were carried out using two different techniques of conjugation; filter and broth mating techniques. The transconjugtion frequencies of the filter mating technique (4.6×10-5) was higher than the broth mating technique (2.4×10-5). The genetic variability among the transcojugants lines were tested using RAPD analysis and showed 13.76% polymorphism percentages for donor, recipient and tranconjugants lines The genes encoding proteins involved in bacteriocin production were isolated and sequenced from E . faecium AH2 (entA, entI, entF, entR , orfA2 , orfA3), Pediococcus pentoceseus AH1( PapA, PedB) and nis A from Lactococcus lactis sub lactis. All sequences of genes were deposited in the GenBank database under accession numbers: LC064146LC064146LC064146 , LC064147LC064147LC064147 , LC064148, LC064149, LC0641LC0641LC0641 50 , LC064151,LC064151,LC064151, LC101300LC101300LC101300 ,LC101489LC101489LC101489and LC101789.and101789.
2016
Ph.d
Cairo
Agriculture