Identification of Lactobacillus plantarum Isolated from Olive by PCR-RFLP

Olive and its products are rich in lactic acid bacteria (LAB). Natural fermentation and bitterness process of olive is performed by lactic acid bacteria especially Lactobabacillus and in particular, Lactobacillus plantarum which is one of the probiotic bacteria. Today, Biochemical Approaches and molecular tools such as PCR-based methods are alternative methods commonly used for bacterial identification. The aim of this study was biochemical and molecular identification of Lactobacillus plantarum among lactic acid bacteria isolated from Iranian native olive. In this research, 28 LAB strains isolated from different kinds of Iranian native olive and standard strain of Lactobacillus plantarum were cultured on MRS-agar and biochemical identification tests such as carbohydrate fermentation were performed on each strain. Bacterial DNA extraction was performed by phenol-chloroform method. Specific primers were designed for 16SrDNA+ITS sequence of Lactobacillus plantarum and PCR was performed by touch-up PCR method. In order to more evaluation of the strains and identification the differences between them, the RFLP technique was used and the effects of TaqI and HaeIII enzymes were analyzed on PCR products. From 28 isolated strains, 5 strains were formed strong band by specific primers of Lactobacillus plantarum. One of the PCR products was sent for sequencing and resulting data was confirmed that it was Lactobacillus plantarum. There is no differences between RFLP results on the level of two enzymes, HaeIII and TaqI, but carbohydrate pattern of one strain was slightly different. The result indicated that Lactobacillus plantarum could be isolated from Iranian native olive. According to natural presence of these bacteria, by enrichment and increasing the number of them during fermentation and bitterness stages, the probiotic value of olive could be increased.