Evaluation of the rrn operon copy number in Bifidobacterium using real-time PCR

• Published in: Letters in applied microbiology Vol. 38; no. 3; pp. 229 - 232
• Main Authors: Candela, M, Vitali, B, Matteuzzi, D, Brigidi, P
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Science Ltd 01.01.2004; Blackwell Science
• Subjects: Bifidobacterium; Bifidobacterium - genetics; Biological and medical sciences; Blotting, Southern; DNA, Bacterial - analysis; DNA, Bacterial - isolation & purification; DNA, Ribosomal - analysis; DNA, Ribosomal - isolation & purification; Fundamental and applied biological sciences. Psychology; Gene Dosage; gene quantification; Microbiology; Polymerase Chain Reaction - methods; Polymorphism, Genetic; probiotics; real‐time PCR; ribosomal operon redundancy; RNA, Ribosomal, 16S - genetics; rRNA Operon - genetics; ribosomal operon redundancy; Bifidobacterium; Actinomycetaceae; Applied microbiology; Copy number; Operon; real-time PCR; Real time; gene quantification; Actinomycetales; Polymerase chain reaction; Bacteria; Actinomycetes; probiotics
• ISSN: 0266-8254; 1472-765X; 1365-2673
• DOI: 10.1111/j.1472-765x.2003.01475.x

Aims:  A real‐time PCR‐based method was developed to evaluate the Bifidobacterium rRNA operon copy number. As a result of their repetitive nature, rRNA operons are very suitable targets for chromosomal integration of heterologous genes. Methods and Results:  The rrn operon multiplicity per chromosome was determined by real‐time PCR quantification of the 16S rRNA amplicons obtained from genomic DNA. The values obtained in several bifidobacterial strains of human origin ranged from 1 to 5. The reliability of the method developed was confirmed by Southern hybridization technique. Conclusions:  In the Bifidobacterium genus the rrn operon copies showed variability at species and strain level. The identification of Bifidobacterium strains with high rRNA multiplicity allowed the selection of potential hosts for chromosomal integration. Significance and Impact of the Study:  The methodology here proposed represents a rapid, reliable and sensitive new tool for the quantification of rrn operon copy number in bacteria.