Determination of DNA Transfer Frequency, Related to the Presence of comX Gene, in Natural Streptococcus thermophilus Isolates

Abstract

Activity of comX, alternative sigma factor, is crucial in the expression of the genes required for conjugation takes place in Streptococci and hence it is related to the frequency of competency. Determination of these genes in natural isolates, therefore, is a pivotal issue to understand the conjugation capacity of those microorganisms. In the current study, 106 Streptococcus thermophilus isolates are investigated for the presence of comX gene. Total 46 of these isolates are found to be comX positive using polymerase chain reaction (PCR) and nucleotide sequencing data of them were obtained. The plasmid pNZ276 and catalase tests were used to investigate the conjugation frequency of competent cells. Conjugation characteristics of the isolates BioSt-7, 16, 19, 23, 24, 25, 29, 30, 36, 37, 42 and 46 were studied and conjugation frequency of the isolates BioSt-19, 24, 25,29 and 42 are found to be over 40% and moreover a notable relationship between conjugation ratio and comX gene is determined. Substitution of isoleucine (I) for asparagine (N) in 72nd amino acid position of comX protein increased the conjugation frequency of the isolates. The findings of current study suggest that, some mutations occurred on comX gene may affect the frequency of competency and conjugated characteristics of streptococcal isolates.

Keywords

• Streptococcus thermophilus
• comX gene
• conjugation frequency

Doğal Streptococcus thermophilus İzolatlarında comX Gen Varlığına Bağlı DNA Transfer Frekansının Belirlenmesi

Abstract

Alternatif sigma faktör olan comX, streptokoklarda dışarıdan DNA alabilme kapasitesini belirleyen genlerin transkripsiyonunu kontrol eden ve dolaysıyla bu suşların yabancı genetik materyali kabul etme frekansı ile ilişkili olan bir gendir. Bu nedenle doğal izolatlardaki varlığının tespit edilmesi, bu izolatların konjugasyon kapasitelerinin belirlenmesi açısından önem arz etmektedir. Bu çalışmada 106 farklı Streptococcus thermophilus izolatında comX geninin varlığı araştırılmıştır. Bu izolatların 46 tanesinde polimeraz zincir tepkimesi (PZT) yöntemi ile comX gen varlığı tespit edilmiş ve nükleotid dizileri belirlenmiştir. Kompetent hücrelerin konjugasyon frekansları için pNZ276 plazmiti ve katalaz testi kullanılmıştır. BioSt-7, 16, 19, 23, 24, 25, 29, 30, 36, 37, 42 ve 46 nolu izolatların konjugasyon özellikleri tespit edilmiştir. BioSt-19, 24, 25,29 ve 42 nolu izolatların da konjugasyon frekansları % 40’tan yüksek bulunmuş ve konjugasyon oranı ile comX gen dizisi arasında ilişki tespit edilmiştir. ComX proteininde 72. amino asit pozisyonundaki asparajinin yerine izolösin değişimi (N→I), konjugasyon frekansını artırmıştır. Bu çalışmanın sonuçları, comX gen dizisi üzerindeki değişim mutasyonlarının izolatların gen transfer sıklığını önemli derecede etkileyebileceğini, suşlara göre farklılık içereceğini ve konjugasyon özelliğini değiştirebileceğini göstermiştir.

Keywords

• Streptococcus thermophilus
• comX geni
• konjugasyon frekansı

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