API-ZYM and numerical analysis of 16S rRNA gene ıdendified micromonospora ısolates from the Black Sea region

Abstract

In this study, molecular and numerical methods were applied to identify possible novel Micromonospora isolates. The isolation study of different actinomycetes obtained from the Black Sea Region soil samples were carried out by modifying different isolation techniques. Genomic DNA of bacteria is obtained by using DNA isolation kit. PCR-mediated amplification of the 16S rRNA gene region of test isolates were performed by using 27F and 1525R universal primers. Almost complete 16S rRNA gene sequences of the isolates were determined using automatic sequencer with 800R, MG3F and MG5F primers. Phylogenetic trees based on 16S rRNA gene were constructed with the neighbour-joining, maximum likelihood and maximum parsimony algorithms by using MEGA6 software. In this study, we obtained one hundred fifty-one isolates in total, and it was determined that these microorganims belong to different groups of actinomycetes. Eleven possible novel strains which belong to the genus Micromonospora isolates were subjected to numerical and biochemical analysis. Putative novel Micromonospora soil isolates are expected to contribute to taxonomy of actinomycetes and also many novel species are sure to emerge

Keywords

• actinomycetes,Micromonospora,16S rRNA gene sequencing,numeric taxonomy

Karadeniz bölgesinden izole edilen 16S rRNA gen bölgesi ile tanımlanan Micromonospora izolatlarının APIZYM ve nümerik analizleri

Abstract

Çalışmamızda yeni tür olması muhtemel Micromonospora izolatlarına moleküler ve nümerik metodlar uygulanmıştır. Karadeniz bölgesinden alınan toprak örneklerinden izole edilen farklı aktinomisetlerin izolasyon çalışması farklı izolasyon teknikleri kullanılarak gerçekleştirilmiştir. Bakterilerin genomic DNA’sı DNA izolasyon kiti kullanılarak elde edilmiştir. Test izolatlarına ait 16S rRNA gen bölgesi çoğaltımı 27F ve 1525R primerleri ile gerçekleştirilmiştir. İzolatlara ait gen dizileri 800R, MG3F ve MG5F evrensel primerleri ile otomatik dizileme cihazı kullanılarak belirlenmiştir. 16S rRNA gen bölgesi temelli filogenetik ağaçlar MEGA6 programında neighbour-joining, maximum likelihood ve maximum parsimony algorithmaları kullanılarak oluşturulmuştur. Bu çalışmada, toplamda yüz elli bir aktinomiset izolatı elde edilmiş ve bunların farklı aktinomiset gruplarına dahil oldukları belirlenmiştir. Bu izolatlardan Micromonospora cinsine ait olan yeni tür olması muhtemel 11 tanesine nümerik ve biyokimyasal testler uygulanmıştır. Yeni olduğu varsayılan Micromonospora toprak izolatlarının aktinomiset taksonomisine katkı sağlayacağı ve aynı zamanda birçok yeni türün ortaya çıkacağı düşünülmektedir

Keywords

• aktinomiset,Micromonospora,16S rRNA gen sekansı,nümerik taksonomi

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