The use of qualitative and quantitative polymerase chain reactions for diagnosis of cytomegalovirus infections in bone marrow and kidney transplant recipients

Marin, Lauro Juliano; Cunha, Aldo Albuquerque; Aquino, Víctor Hugo; Figueiredo, Luíz Tadeu Moraes

doi:10.1590/s0037-86822004000200009

Cited by 3 publications

(4 citation statements)

References 19 publications

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“…For example, among AIDS patients, gB2 has been significantly associated with retinitis in some reports, unlike other studies [33]. Among BMT patients, gB3 and gB4 were associated with higher mortality by myelosuppression [20,16] and were infrequently associated with graft-versus-host disease.…”

Section: Discussionmentioning

confidence: 98%

Clinical correlations of human cytomegalovirus strains and viral load in kidney transplant recipients

Nogueira

Ozaki

Tomiyama

et al. 2009

International Immunopharmacology

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Section: Discussionmentioning

confidence: 98%

Clinical correlations of human cytomegalovirus strains and viral load in kidney transplant recipients

Nogueira

Ozaki

Tomiyama

et al. 2009

International Immunopharmacology

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“…Underwood et al (1994) were the first to propose the determination of the number of genes by comparing samples with a standard curve, plotted based on DNA amplification of samples with previously determined gene dosages. In the last years, semiquantification works have been carried out in a similar way in a wide range of cases such as in the investigation of human diseases (diagnosis and determination of cytomegalovirus load in the leukocytes of bone marrow or kidney transplant patients (Marin et al, 2004) and determination of interferon-gamma mRNA levels in AIDS-free HIV-infected individuals (Barabás et al, 2001)), the identification of mixtures containing conventional and genetically modified maize seeds (Tozzini et al, 2000), or the determination of transgenic maize and soybean composition in processed foods (Cazzola and Petrucelli, 2006). However, its longest and most frequent use is in the analysis of gene expression, that is SQPCR, for instance, in the study carried out by Chelly et al (1990) concerning the expression analysis of the gene for muscular dystrophy, or in the distinction between queen and worker bees according to the production of mRNA for the enzyme methyltransferase (Baffi et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

“…Although SQPCR does not permit an absolute quantification, the above-mentioned works demonstrated that it was possible to estimate the relative amount (Chelly et al, 1990;Baffi et al, 2004;Dutta et al, 2007;Tao et al, 2007) or the number of amplified molecules in each sample (Underwood et al, 1994;Tozzini et al, 2000;Marin et al, 2004;Cazzola and Petrucelli, 2006). The sensitivity of SQPCR is the result of a chain reaction, in which the products from one amplification cycle serve as substrates for the next, resulting in an exponential increase of the product.…”

Section: Introductionmentioning

confidence: 99%

Distinction between plant samples according to allele dosage by semiquantitative polymerase chain reaction

Martins¹,

Carneiro²,

Guimarães³

et al. 2009

Genet. Mol. Res.

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AbSTrACT. The lack of informativity of samples from heterozygotic individuals is one of the hindrances in the mapping of quantitative trait loci of outbred populations, since it is not normally possible to identify the origin of each allele. One way to include these individuals in analyses would be to genotype their endosperm, considering that a heterozygote (Aa) has AAa or Aaa endosperm, when the female genitor donates the A or a allele, respectively. We used semiquantitative polymerase chain reaction to determine allele dosages in DNA mixtures, by simulating the observed conditions for endospermic tissue. Semiquantitative polymerase chain reaction on agarose gels, along with regression analysis, allowed differentiation of the samples according to the amount of DNA. This type of information will help decrease the number of non-informative individuals in quantitative trait locus mapping of outbred populations, thereby increasing mapping accuracy.

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“…We are indebted to Klaus Roemer for critical review of the manuscript. GAA ACG CGC G C GC AAT CGG 81873-81890 2, 3, 9, 12, 22 R2 (9,12,22) TAC GCT GCA GTT CAC/CCC AG 82055-82036 2 R3 (11) T-G AAC TGG AAC GTT TGG C 82174-82156 2, 8, 9, 12, 13, 22 F4 (18, 19) CGG AAA CGA TGG TGT AGT TGG 82571-82591 R5 (18,19) TCC AAC ACC CAC T/AG ACC GGT 82838-82818 F6 (15) ATA GGA GGC GCC ACG TAT TC …”

mentioning

confidence: 99%

Errors in Published Sequences of Human Cytomegalovirus Primers and Probes: Do We Need More Quality Control?

2005

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Over the past decade, many authors have focused on PCR as a powerful technique for the evaluation of human cytomegalovirus (CMV). The key to PCR lies in the design of oligonucleotides, as the specific sequences largely affect PCR's efficacy and sensitivity. This study was designed to examine the quality of published sequences of CMV primers and probes.PubMed was searched in the National Center for Biotechnology Information website (http://www.ncbi.nlm.nih.gov) for English peer-reviewed articles using CMV and PCR as keywords. Articles reporting on virus genotyping or species-level identification, as well as letters to editors and reviews were excluded. The full texts of 91 papers published between 1993 and 2004 were studied. Of these, 34 papers did not describe the detailed nucleotide sequence, including 17 papers using commercially available kits. The remaining 57 papers with a total of 199 CMV-specific oligonucleotides were examined. Oligonucleotides with identical sequences or with one additional nucleotide at either the 3Ј or 5Ј end of the sequence were identified as synonymous.Using The Sequence Manipulation Suite web-based programs (written by Paul Stothard, University of Alberta, Canada), the binding sites of all 199 oligonucleotides were identified using GenBank strain AD169 genome sequence (GenBank accession no. X17403 and NC_001347). Mismatches to all published sequences of CMV were analyzed by the Basic Local Alignment Search Tool (BLAST) (http://www .ncbi.nlm.nih.gov/BLAST/). Papers containing oligonucleotide errors were studied for technical reasons for employment of mutated oligonucleotides or subsequently published errata. Moreover, information on identified errors was conveyed to all corresponding authors and coauthors.Ten oligonucleotides did not match any CMV strain, and reasons were specified neither in the respective articles nor in authors' replies (Table 1). Primers R2, F6, and F9 were incorrectly transferred from previous publications (2, 5, 7) as indicated by the authors (V. H. Aquino and X. L. Pang, personal communication). The oligonucleotides F1, R3, and P10 were apparently also incorrectly transferred from prior publications (14). Moreover, primers F4 and R5 possessed mismatches at their 3Ј-end triplets, which may reduce PCR efficiency (21). Furthermore, the two degenerated probes (P7 and P8) contained Y (C/T) instead of R (A/G) in P7 and vice versa in P8. Obviously, these mismatches may drastically affect the efficacies of these probes (17).In summary, we show that 5% of the CMV oligonucleotides included mismatches to all published sequences half of which were incorrectly reported mostly in secondary publications. Moreover, since we considered all sequences correct when they matched at least one CMV strain, even when the authors used a different strain for design or admitted an error, the number of errors is underestimated by our approach. These observations point to the possibility that the report of erroneous primers and probes is a widespread problem irritating other researchers. I...

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The use of qualitative and quantitative polymerase chain reactions for diagnosis of cytomegalovirus infections in bone marrow and kidney transplant recipients

Abstract: The purpose of this work was to test a cytomegalovirus qualitative PCR and a semi-quantitative PCR on the determination of CMV load in leukocytes of

Cited by 3 publications

References 19 publications

Clinical correlations of human cytomegalovirus strains and viral load in kidney transplant recipients

Clinical correlations of human cytomegalovirus strains and viral load in kidney transplant recipients

Distinction between plant samples according to allele dosage by semiquantitative polymerase chain reaction

Errors in Published Sequences of Human Cytomegalovirus Primers and Probes: Do We Need More Quality Control?

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