A Method to Compensate for Different Amplification Efficiencies

Ogino, Shuji; Meijerink, Jules P.P.

doi:10.1016/s1525-1578(10)60671-x

Cited by 3 publications

(3 citation statements)

References 6 publications

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“…In this context, the internal standard approach has proven its suitability and, accordingly, has found a wide range of applications for environmental analyses (Luciano et al 2007;Mas et al 2008). Amplification with the same primer pair and thus with the same efficiencies as the target internal standards is therefore most accurate (Ogino 2001). Even if the PCR was inhibited by about 70%, the application of an adapted internal standard method allowed to correctly quantify DNA copy numbers (Stocher and Berg 2002).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Methanogenic population dynamics during semi‐continuous biogas fermentation and acidification by overloading

Blume

Bergmann

Nettmann

et al. 2010

Journal of Applied Microbiology

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Aims: This study was to investigate the methanogenic community in a biogas reactor from start‐up to acidification conditions. Furthermore, reliability and accuracy of the applied quantitative real‐time PCR method (Q‐PCR) was briefly evaluated. Methods and Results: A mesophilic (37°C), maize silage fed, continuously stirred tank reactor was surveyed. It was operated semi‐continuously with increasing daily organic loading rates (OLRs) to reach acidification. Gas production and organic acid composition were measured. Methanogenic community structure was determined by 16S rDNA‐based Q‐PCR to estimate the abundance of key methanogenic micro‐organisms. 16S rDNA of hydrogenotrophic Methanobacteriales was most abundant at OLRs of ≥3·7 g dry organic matter (DOM) l−1 day−1. By contrast, that of aceticlastic Methanosaetaceae predominated at lower OLRs but disappeared at OLRs of ≥4·1 g DOM l−1 day−1. At the same OLR, the propionate concentration increased dramatically indicating the acidification of the digester. Application of internal standards to examine Q‐PCR’s accuracy revealed that the detected amount of 16S rDNA may vary within one log cycle. Conclusions: These results suggest that the absence of Methanosaetaceae might be taken as biological indicator for process’ instability. Inhibitory effects on Q‐PCR analyses could not be determined based on the spiking experiments. Significance and Impact of the Study: In this study, reactors’ microbiology was observed over time using Q‐PCR. Insights into the abundance of different methanogens might be used to improve the performance of biogas reactors.

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Section: Introductionmentioning

confidence: 99%

“…2008). Amplification with the same primer pair and thus with the same efficiencies as the target internal standards is therefore most accurate (Ogino 2001). Even if the PCR was inhibited by about 70%, the application of an adapted internal standard method allowed to correctly quantify DNA copy numbers (Stocher and Berg 2002).…”

Section: Introductionmentioning

confidence: 99%

Methanogenic population dynamics during semi‐continuous biogas fermentation and acidification by overloading

Blume

Bergmann

Nettmann

et al. 2010

Journal of Applied Microbiology

View full text Add to dashboard Cite

show abstract

“…When comparing the two separate assays the efficiency should not vary more than 5% (Ogino, 2001). The efficiencies of the MqPCR assay were comparable if not greater than the S-qPCR assay.…”

Section: Discussionmentioning

confidence: 94%