Selection of specific primers based on 16s rRNA gene for Bacillus cereus bacteria

ABSTRACT The bacterial candidate phylum Termite Group I (TG-1) presently consists mostly of “Endomicrobia,” which are endosymbionts of flagellate protists occurring exclusively in the hindguts of termites and wood-feeding cockroaches. Here, we show that public databases contain many, mostly undocumented 16S rRNA gene sequences from other habitats that are affiliated with the TG-1 phylum but are only distantly related to “Endomicrobia.” Phylogenetic analysis of the expanded data set revealed several diverse and deeply branching lineages comprising clones from many different habitats. In addition, we designed specific primers to explore the diversity and environmental distribution of bacteria in the TG-1 phylum.

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Overestimation of Streptococcus mutans prevalence by nested PCR detection of the 16S rRNA gene

Journal of Medical Microbiology ◽

10.1099/jmm.0.46280-0 ◽

2006 ◽

Vol 55 (1) ◽

pp. 109-113 ◽

Cited By ~ 14

Author(s):

Ali Al-Ahmad ◽

Thorsten Mathias Auschill ◽

Gabriele Braun ◽

Elmar Hellwig ◽

Nicole Birgit Arweiler

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Streptococcus Mutans ◽

Nested Pcr ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Direct Pcr ◽

Specific Primers ◽

The 16S Rrna Gene

This study was carried out in order to compare two PCR-based methods in the detection of Streptococcus mutans. The first PCR method was based on primers for the 16S rRNA gene and the second method was based on specific primers that targeted the glucosyltransferase gene (gtfB). Each PCR was performed with eight different streptococci from the viridans group, five other streptococci and 17 different non-streptococcal bacterial strains. Direct use of the S. mutans 16S rRNA gene-specific primers revealed that Streptococcus gordonii and Streptococcus infantis were also detected. After amplifying the 16S rRNA gene with universal primers and subsequently performing nested PCR, the S. mutans-specific nested primers based on the 16S rRNA gene detected all tested streptococci. There was no cross-reaction of the gtfB primers after direct PCR. Our results indicate that direct PCR and nested PCR based on 16S rRNA genes can reveal false-positive results for oral streptococci and lead to an overestimation of the prevalence of S. mutans with regards to its role as the most prevalent causative agent of dental caries.

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Genus-specific primers targeting the 16S rRNA gene for PCR detection of members of the genus Verrucosispora

Antonie van Leeuwenhoek ◽

10.1007/s10482-011-9571-4 ◽

2011 ◽

Vol 100 (1) ◽

pp. 117-128 ◽

Cited By ~ 6

Author(s):

Qingyi Xie ◽

Kui Hong ◽

Michael Goodfellow

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Pcr Detection ◽

Rrna Gene ◽

Specific Primers ◽

The 16S Rrna Gene

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KAJIAN MOLEKULER PADA PROBIOTIK ASAL AIR SUSU IBU DALAM SINTESIS EKSOPOLISAKARIDA (EPS)

Jurnal Bioteknologi & Biosains Indonesia (JBBI) ◽

10.29122/jbbi.v8i1.4554 ◽

2021 ◽

Vol 8 (1) ◽

pp. 114-123

Author(s):

Hamiyawati Qoimatu Dini Alfaruqi ◽

Nosa Septiana Anindita ◽

Arif Bimantara

Keyword(s):

Breast Milk ◽

16S Rrna ◽

16S Rrna Gene ◽

Universal Primers ◽

Rrna Gene ◽

Human Breast Milk ◽

Human Breast ◽

Adhesion Ability ◽

The 16S Rrna Gene ◽

Selection Of

Molecular Studies on Probiotic of Human Breast Milk in the Synthesis of Exopolysaccharide (EPS) The glucosyltransferase (gtf) gene has an important role in exopolysaccharide (EPS) synthesis in probiotic bacteria. The EPS produced is associated with the adhesion ability of bacteria to the intestinal mucosa. Therefore, the gtf gene can be used as a parameter in the selection of potential probiotic through a molecular approach. This study was conducted to determine the presence of the gtf gene in probiotic from human breast milk using PCR technique. The methods in this study include the following: reculture of probiotic isolates, DNA isolation, amplification of the 16S rRNA gene using universal primers (pA and pB), amplification of specific LAB primers (LABfw and LABrv), specific primary design for the gtf gene, and the amplification of the gtf gene. The results of 16S rRNA gene amplification using universal primers obtained the amplicons of 500-1,000 bp in size. The results of amplification using specific LAB primers obtained an amplicon of about 700 bp in all isolates. The results of amplification of the gtf gene using a specific primer produced an amplicon of 325 bp in all isolates. Based on this study, it was concluded that 16 probiotic isolates from human breast milk were proven to have the gtf gene. Gen glukosiltransferase (gtf) memiliki peran penting dalam sintesis eksopolisakarida (EPS) pada bakteri probiotik. EPS yang diproduksi berhubungan dengan kemampuan adhesi bakteri pada mukosa usus. Oleh karena itu, gen gtf dapat dijadikan sebagai salah satu parameter dalam seleksi probiotik potensial melalui pendekatan molekuler. Penelitian ini dilakukan untuk mengetahui adanya gen gtf pada probiotik asal air susu ibu (ASI) menggunakan teknik PCR. Metode pada penelitian ini meliputi: reculture isolat probiotik, isolasi DNA, amplifikasi gen 16S rRNA menggunakan primer universal (pA dan pB), amplifikasi primer spesifik BAL (LABfw dan LABrv), desain primer spesifik untuk gen gtf dan amplifikasi gen gtf. Hasil amplifikasi gen 16S rRNA menggunakan primer universal diperoleh amplikon berukuran antara 500-1.000 bp. Adapun hasil amplifikasi menggunakan primer spesifik BAL diperoleh amplikon berukuran sekitar 700 bp pada seluruh isolat. Hasil amplifikasi gen gtf menggunakan primer spesifik menghasilkan amplikon berukuran sekitar 325 bp pada seluruh isolat. Berdasarkan penelitian ini dapat disimpulkan bahwa 16 isolat probiotik asal ASI terbukti memiliki gen gtf.

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Identification and Assessment of Genetic Similarity of Soil Bacterial Isolates of Pseudomonas spp. Using Molecular Techniques

Polish Journal of Microbiology ◽

10.33073/pjm-2014-039 ◽

2014 ◽

Vol 63 (3) ◽

pp. 291-298

Author(s):

ANNA LISEK ◽

LIDIA SAS PASZ ◽

PAWEŁ TRZCIŃSKI

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Genetic Similarity ◽

Sour Cherry ◽

23S Rrna ◽

Rrna Gene ◽

Bacterial Isolates ◽

Bacterial Strains ◽

The 16S Rrna Gene ◽

Selection Of

Bacteria of the genus Pseudomonas are often components of bioproducts designed to enhance the condition of the soil and plants. The use of Pseudomonas bacteria in bioproducts must be preceded by the acquisition, characterization and selection of beneficial strains living in the soil. A prerequisite for the selection of bacterial strains for use in bioproducts is to be able to identify the isolates rapidly and accurately. To identify and differentiate 15 bacterial isolates obtained from the soil surrounding the roots of sour cherry trees and to assess their genetic similarity, the rep-PCR technique and restriction analysis of the 16S rRNA gene and the 16S-ITS-23S rRNA operon were used. In addition, a sequence analysis of the 16S rRNA gene was performed. The analyses made it possible to divide the isolates into four clusters and to confirm their affiliation with the Pseudomonas species. RFLP analysis of the 16S-ITS-23S rRNA operon enabled greater differentiation of the isolates than RFLP of the 16S rRNA gene. The greatest differentiation of isolates within the clusters was obtained after using the rep-PCR technique. However, none of the techniques was able to discriminate all the isolates, which indicates very high genetic similarity of the Pseudomonas isolates found in the same sample of soil from around the roots of sour cherry trees. The tests performed will find application for distinguishing and identifying Pseudomonas strains collected from the soil in order to select the most valuable bacterial strains that produce beneficial effects on plants.

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