An improved, real-time PCR assay for the detection of GC-rich and low abundance templates of Mycobacterium tuberculosis.
The detection of low abundance mRNA and/or GC-rich targets is very difficult using real-time PCR, often requiring laborious optimization procedures. This work shows that formamide is a useful PCR additive, increasing the sensitivity and specificity of SYBR Green real-time PCR to detect low abundance mycobacterial RNA from infected samples.
Institute Neurobiology and Molecular Medicine, Molec. Med. Section, National Research Council, Via del Fosso del Cavaliere, 100, 00133, Rome, Italy.
Manuela
M
Grassi
Elisabetta
E
Volpe
Vittorio
V
Colizzi
Francesca
F
Mariani
D001482:Base Composition; D052160:Benzothiazoles; D003959:Diamines; D005456:Fluorescent Dyes / Q000737:chemistry; D005559:Formamides / Q000737:chemistry; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000302:isolation & purification; D009930:Organic Chemicals / Q000737:chemistry; D011804:Quinolines; D012333:RNA, Messenger / Q000032:analysis* / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013698:Templates, Genetic; D014376:Tuberculosis / Q000382:microbiology
D052160:Benzothiazoles; D003959:Diamines; D005456:Fluorescent Dyes; D005559:Formamides; D009930:Organic Chemicals; D011804:Quinolines; D012333:RNA, Messenger; C098022:SYBR Green I; C031066:formamide