Rapid techniques for DNA extraction from routinely processed archival tissue for use in PCR.

Sepp, Róbert; Szabó, Ildikò; Uda, Hirotsugu; Sakamoto, Haruhiko

doi:10.1136/jcp.47.4.318

Cited by 147 publications

(116 citation statements)

References 17 publications

(6 reference statements)

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“…Also, the reproducibility of extracting amplifiable DNA was rather low . In part, this may be explained by the presence of excessive staining residues (Gall et al, 1993;Sepp et al, 1994). Still, we observed in this study that the DNA integrity hardly improved after an additional purification step (salting-out procedure) following proteinase K treatment.…”

Section: Discussionmentioning

confidence: 50%

A simplified and reliable HPV testing of archival Papanicolaou-stained cervical smears: application to cervical smears from cancer patients starting with cytologically normal smears

et al. 2000

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SummaryThe efficacy of four methods to recover DNA from Papanicolaou (Pap)-stained archival cervical smears for optimal detection of human papillomavirus (HPV) DNA by GP5+/bioGP6+ polymerase chain reaction (PCR) was investigated. Two of the methods were based on proteinase K treatment and two based on treatment with guanidinium thiocyanate (GTC). The quality of the DNA as measured by PCR assays amplifying different sizes of the β-globin gene appeared to be superior for the GTC-based assays. Using competitive β-globin PCR assays, one of the GTC-based, assays, provisionally named High Pure PCR Template Preparation (HPPTP) assay, yielded by far the highest quantity of amplifiable DNA. It allowed the recovery of 2.2 × 10 5 to 3 × 10 5 genome equivalents in smears containing 5 × 10 5 to 20 × 10 5 nucleated cells, indicating a mean efficiency of 26% (range of 15-44%). In contrast, the other methods revealed markedly lower efficiencies varying from 1% to 10%. The use of the HPPTP assay as a reliable processing procedure was validated by demonstrating a complete agreement in HPV detection and 93% agreement in HPV typing between 39 archival Pap-stained and paired fresh-frozen cervical smears. This method was applied to 40 archival smears from ten cervical cancer patients (selected from a group of 200 patients) which had a history of 3-6 smears with the first smear being Pap 1 or 2 taken at least 5 years before cancer was diagnosed. The average time period between the first Pap 1/2 smear that contained the same HPV type as in the corresponding carcinoma and diagnosis of cervical cancer was 12.0 ± 2.9 years. All subsequent smears were invariably positive for the same HPV type which was also found in the cervical cancer biopsy. In conclusion, the HPPTP assay provides a reliable and efficient means to extract DNA from Pap-stained archival cervical smears for the detection of HPV DNA by PCR and would be the method of choice for future HPV analysis of archival Pap-stained cervical smears.

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Section: Discussionmentioning

confidence: 50%

A simplified and reliable HPV testing of archival Papanicolaou-stained cervical smears: application to cervical smears from cancer patients starting with cytologically normal smears

et al. 2000

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“…Then, genomic DNA was purified by phenol/ chloroform extraction. Genomic DNA of some samples was purified from parafilm-embedded blocks (37). The surgical tissues were thinsliced, and parafilm was removed with sufficient amount of xylene.…”

Section: Methodsmentioning

confidence: 99%

The −251T Allele of the Interleukin-8 Promoter Is Associated with Increased Risk of Gastric Carcinoma Featuring Diffuse-Type Histopathology in Chinese Population

Lee

Tai

Lan

et al. 2005

Clinical Cancer Research

139

100

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Purpose: Persistent interleukin-8 (IL-8) production contributes to chronic inflammation of the stomach. The proinflammatory IL-1h polymorphisms, which enhance the cytokine production, are associated with increased risk of gastric cancer. The À251A/T polymorphism of the IL-8 promoter is involved in several human diseases. Particularly, the À251A is associated with decreased risk of colorectal cancer. We aimed to determine whether the À251 allele resulting in high IL-8 expression was associated with increased risk of gastric carcinoma. Experimental Design: The À251A/T promoters were cloned and analyzed by luciferase assay. Binding of nuclear proteins to the À251A/T promoters was analyzed by electrophoretic mobility shift assay.The À251A/T promoters were differentiated by PCR-RFLP. Comparison of gastric cancer risk between the À251A/T promoters was done by a case-control study. Results:The À251Tallele possessed transcriptional activity 2-to 5-fold stronger than the À251A counterpart. Electrophoretic mobility shift assay showed that the À251A promoter had strong ability to bind to an unknown protein or multiprotein complex. The À251T allele was associated with increased risk of noncardia (P trend = 0.012) and cardia (P trend = 0.029) carcinomas. Gastric carcinoma patients with the low-risk AA genotype had a tendency to sustain intestinal-type carcinomas (m 2 = 6.816; P = 0.033); however, the high-risk À251Tallele was associated with >2-fold increased risk of diffuse-type (AA versus AT + TT: odds ratio, 2.52; 95% confidence interval,1.16-5.49; P = 0.017) and mixed-type (AA versus AT + TT: odds ratio, 2.22; 95% confidence interval, 1.12-4.40; P = 0.019) carcinomas. Conclusions:The IL-8 À251Tallele is significantly associated with increased risk of gastric carcinoma, particularly the diffuse and mixed types in Chinese population.

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“…Moreover, Chelex-100 chelates polyvalent metal ions, which may act in the breakdown of DNA and PCR inhibition (Sepp et al, 1994;Barea et al, 2004). As this protocol only requires a low sample volume (25 μL) (Manuja et al, 2006), it can be applicable to situations where the quantity of biological material is limited, for example, in forensic assays (Koch and Andrade, 2008;Walsh et al, 1991).…”

Section: Discussionmentioning

confidence: 99%

“…However, it is clear that the use of the DNeasy Blood & Tissue Kit is limited by its high cost (Bailes et al, 2007), and extraction with phenol-chloroform is a laborious and potentially hazardous method (Sepp et al, 1994), where a significant amount of DNA can be lost (Goldenberger et al, 1995) and degraded (Hossain et al, 1997) and the PCR inhibited (Goldenberger et al, 1995). By contrast, Chelex-100 resin emerged as a cheap, effective, fast, and simple method, which can be realized in few steps, and does not require the use of organic solvents, which confirms observations of previous studies (Walsh et al, 1991;Simonato et al, 2007).…”

Section: Discussionmentioning

confidence: 99%

Comparative study of DNA extraction methodologies from goat sperm and its effects on polymerase chain reaction analysis

Silva¹,

Pelinca²,

Acosta³

et al. 2014

Genet. Mol. Res.

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ABSTRACT. Successful DNA extraction is indispensable for molecular methods based on polymerase chain reaction (PCR); however, goat sperm DNA extraction is limited. Thus, the aim of this study was to evaluate three methods to extract DNA from goat sperm for use in PCR. Eight goat semen pools were used for DNA extraction by using the DNeasy Blood & Tissue Kit, phenol-chloroform, and Chelex-100 methods. DNA samples were analyzed spectrophotometrically to determine the DNA concentration and purity, visualized on 0.8% agarose gel, and used at different amounts (150, 100, 50, 10, and 1 ng) for PCR with electrophoresis, followed by 1.5% agarose gel electrophoresis. The quantity of DNA extracted with Chelex-100 was DNA extraction from goat sperm for PCR analysis higher (P < 0.05) than that obtained with either the DNeasy Blood & Tissue Kit or the phenol-chloroform method, with the phenolchloroform method yielding a greater quantity (P < 0.05) than the kit. The DNeasy Blood & Tissue Kit produced a higher (P < 0.05) purity product than the Chelex-100 method, and all samples obtained by the three protocols were positive for DNA, as assessed by electrophoresis. All of the different concentrations of DNA produced by these methods were amplified by PCR, although for DNA produced by the phenolchloroform method, PCR was only possible after complementary purification. In conclusion, the Chelex-100 method is cheap, secure, simple, fast, and effective, and is a potential tool for extracting goat sperm DNA without limitations in PCR.

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Rapid techniques for DNA extraction from routinely processed archival tissue for use in PCR.

Cited by 147 publications

References 17 publications

A simplified and reliable HPV testing of archival Papanicolaou-stained cervical smears: application to cervical smears from cancer patients starting with cytologically normal smears

A simplified and reliable HPV testing of archival Papanicolaou-stained cervical smears: application to cervical smears from cancer patients starting with cytologically normal smears

The −251T Allele of the Interleukin-8 Promoter Is Associated with Increased Risk of Gastric Carcinoma Featuring Diffuse-Type Histopathology in Chinese Population

Comparative study of DNA extraction methodologies from goat sperm and its effects on polymerase chain reaction analysis

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