Doğal Fermente Olan Sütteki Lactobacillus Spp.16S rRNA Geninin Kısmi Analizi

Abstract

Purpose: The aim of this research is to analysis 16S rRNA gene sequence of Lactobacillus spp. from natural fermented milk. Material and Methods:The methods that used in this study are total DNA isolation using alkaline lysis, PCR amplification carried out using of specific primers, and identification of amplified DNA using sequencing. The result of PCR process is a nuclotide with approximately 585 bp length which contained V1-V3 variable regions. 16S rRNA gene sequence then compared and aligned with existing 16S rRNA data sequence from GenBank. Results: The conserved region of 16S rRNA gene from nuclotide number 189-358 and 384-445 were 95,9% and 94,9% respectively, and the total conserved region of 16S rRNA gene is 78.8%. From total 7 isolates that isolated from natural fermented milk , 2 isolates were identified as L. plantarum with similarity value 98.28% and 97.78%, the other 5 isolates were identified as L. rhamnosus (2 isolates) with similarity value 98.38% and 97.51%, L. zeae (2 isolates) with similarity value 97.36% and 98.38%, and L. casei (1 isolate) with similarity value 98.20%. Conclusion:These results indicate the test isolates are members of the genus Lactobacillus, but classified in different species. Key Words: Lactobacillus, natural fermentation milk, 16S rRNA gene, partial analysis. ÖZETAmaç: Bu çalışmanın amacı doğal olarak fermente olan sütten Lactobacillus spp. türüne ait 16S rRNA ların gen sekansı yapmaktır. Materyal ve Metod: Kullanılan metodlar; alkali lizis yöntemiyle total DNA izolasyonu, spesifik primerlerle PCR amplifikasyonu ve amplifiye edilmiş DNA ların sekans analizlerinin yapılması şeklindedir. Elde edilen PCR ürünleri yaklaşık 585 bp uzunluğunda olup V1-V3 değişken bölgelerini içermektedir. Sekanslanan 16s rRNA daha sonra gen bankasıyla karşılaştırıldı. Bulgular: 16s rRNA geninin korunmuş bölgeleri; 189-358 ve 384-445 sırasıyla %95,9 ile %94,9 oranında korunurken, 16s rRNA geni totalde %78.8 oranında korunmuştur. Doğal sütün fermentasyonundan izole edilen bu 7 izolattan 2 tanesi L.Plantarum(benzerlik oranları %98.28 ve %97.78 diğer 5 izolattan 2'si L. Rhamnosus (benzerlik oranları (98.38 ve %97.51

Molecular characterization and 16S rRNA sequence analysis of probiotic lactobacillus acidophilus isolated from indigenous Dahi (Yoghurt)

“…showed slight mutation or difference in genetic material (DNA) which might be due to difference in environmental conditions of the region or area of isolation, served as a determinant of bacterial identification at both genus and species level (Clarridge, 2004). Kusmiyati et al (2014) isolated lactobacilli from naturally fermented milk product, after 16S rRNA gene sequence analysis and phylogenetic study he found that similar isolates formed sister clad with their respective reference isolate. This showed that after BLAST, phylogenetic tree construction was proved to be a vital tool for proper identification and also for studying sequence similarities and differences among different strains and spp.…”