geoscience.net logo
+ Resolve Article
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter

+ Translate
+ Subscribe to Site Feed
GeoScience Most Shared ContentMost Shared Content

Incorporation of measurement of DNA integrity into qPCR assays


, : Incorporation of measurement of DNA integrity into qPCR assays. Biotechniques 49(6): 893-897

Optimal accuracy of quantitative PCR (qPCR) requires correction for integrity of the target sequence. Here we combine the mathematics of the Poisson distribution and exponential amplification to show that the frequency of lesions per base (which prevent PCR amplification) can be derived from the slope of the regression line between cycle threshold (Ct) and amplicon length. We found that the amplifiable fraction (AF) of a target can be determined from this frequency and the target length. Experimental results from this method correlated with both the magnitude of a damaging agent and with other measures of DNA damage. Applying the method to a reference sequence, we determined the values for lesions/base in control samples, as well as in the AFs of the target sequence in qPCR samples collected from leukemic patients. The AFs used to calculate the final qPCR result were generally >0.5, but were <0.2 in a few samples, indicating significant degradation. We conclude that DNA damage is not always predictable; quantifying the DNA integrity of a sample and determining the AF of a specific qPCR target will improve the accuracy of qPCR and aid in the interpretation of negative results.

US$19.90

PMID: 21143211

DOI: 10.2144/000113567


Other references

Yadav, M.K.; Redman, J.E.; Leman, L.J.; Alvarez-Gutiérrez, J.M.; Zhang, Y.; Stout, C.David.; Ghadiri, M.Reza., 2005: Structure-based engineering of internal cavities in coiled-coil peptides. Cavities and clefts are frequently important sites of interaction between natural enzymes or receptors and their corresponding substrate or ligand molecules and exemplify the types of molecular surfaces that would facilitate engineering of artific...

Thompson M.D.; Grunow A.M., 2007: The geological case for Avalon off North Africa at 595 Ma. Avalon terranes have commonly been pictured as originating along the Amazonian margin of Gondwana based on Nd isotopic signatures of crystalline rocks and 1.0-1.3 Ga detrital zircons that could have derived from South American source belts. We sug...

Doorduin, J.C., 1974: Soil warming in freesia culture. A discussion on the effect of different soil temperatures on freesia leaf and flower bud initiation, aims and methods of soil warming and some data on fuel consumption.

Sakal, P.; Maluf Filho, F.; Iryia, K.; Moura, E.G.; Tomishigue, T.; Scabbia, A.; Ishioka, S., 1996: An endoscopic technique for resection of small gastrointestinal carcinomas. Gastrointestinal Endoscopy 44(1): 65-68

Massoud, F.; Chertkow, H.; Whitehead, V.; Overbury, O.; Bergman, H., 2002: Word-reading thresholds in Alzheimer disease and mild memory loss: a pilot study. To determine whether subjects with early Alzheimer Disease (AD) or Mild Cognitive Impairment (MCI) would demonstrate significant abnormalities on tests of word reading threshold (WRT), spatial contrast sensitivity (SCS), and color discrimination (...

Elango F.; Lozano J.C., 1979: Sexual seed transmission of xanthomonas manihotis. Transmission of X. manihotis in seeds was studied by the direct immunofluorescence and the enzyme-linked immunosorbent (ELISA) assays. The pathogen was detected in seed embryos but not in other tissues. While both healthy and infected seeds showed...

Kato, M.; Blaner, W.S.; Mertz, J.R.; Das, K.; Kato, K.; Goodman, D.S., 1985: Influence of retinoid nutritional status on cellular retinol- and cellular retinoic acid-binding protein concentrations in various rat tissues. Studies were conducted to explore the effects of differences in retinoid nutritional status and of sex on the tissue distribution and levels of cellular retinol-binding protein (CRBP) and of cellular retinoic acid-binding protein (CRABP) in the ra...

Casado-Vela, J.; Lacal, J.Carlos.; Elortza, F., 2013: Protein chimerism: novel source of protein diversity in humans adds complexity to bottom-up proteomics. Three main molecular mechanisms are considered to contribute expanding the repertoire and diversity of proteins present in living organisms: first, at DNA level (gene polymorphisms and single nucleotide polymorphisms); second, at messenger RNA (pr...

Carter-Su, C.; Kimmich, G.A., 1979: Membrane potentials and sugar transport by ATP-depleted intestinal cells: effect of anion gradients. American Journal of Physiology 237(1): C67-C74

Anonymous, 1980: Sorbitol, mannitol, and xylitol in foods. The use of sorbitol, mannitol, and xylitol as artificial sweeteners in foods, beverages, and food products is discussed in a bibliography of 187 abstracts. Concerns discussed are: mutagenicity, toxicity, teratogenicity, cariogenicity, and the gene...