scholarly journals Rapid preparation of tissue DNA from paraffin-embedded blocks and analysis by polymerase chain reaction.

1993 ◽  
Vol 41 (5) ◽  
pp. 765-768 ◽  
Author(s):  
B F Chen ◽  
S Clejan
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Genetic ◽  
P53 Gene ◽  
Molecular Genetic Analysis ◽  
Chain Reaction ◽  
Rapid Preparation ◽  
Paraffin Block ◽  
Pcr Products ◽  
Embedded Blocks ◽  
Polymerase Chain

We have developed a new, easy, and more rapid method for DNA preparation, which avoids contamination. With this method, manual surgical blade scrapings from precisely targeted areas of paraffin block surfaces, without microtome cutting, were used to obtain tissues from 10 different neoplasms. Our results indicate the feasibility of DNA extraction from the scraped paraffin tissue for molecular genetic analysis. We applied this technique successfully to screen for the presence of human papillomavirus using the polymerase chain reaction (PCR) procedure in cases of endocervical, esophageal, and nasopharyngeal carcinomas, and to examine the expression of p53 gene from prostate and gastric adenocarcinomas. We conclude that this procedure is also suitable for purification of PCR products in analysis of the mutation or loss of allelic genes by Bstu I endonuclease digestion.

Central neurocytoma: morphological, flow cytometric, polymerase chain reaction, fluorescence in situ hybridization, and karyotypic analyses

1999 ◽  
Vol 90 (2) ◽  
pp. 348-354 ◽  
Author(s):  
Venita Jay ◽  
Vern Edwards ◽  
Eelco Hoving ◽  
James Rutka ◽  
Laurence Becker ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
In Situ Hybridization ◽  
Ganglion Cells ◽  
Molecular Genetic ◽  
Central Neurocytoma ◽  
Molecular Genetic Analysis ◽  
Fish Analysis ◽  
Chain Reaction ◽  
Polymerase Chain

✓ The results of cytogenetic and molecular genetic analysis of a central neurocytoma are presented. Central neurocytomas are intriguing neoplasms that exhibit primarily neuronal, but also glial characteristics, which indicate an origin from a pluripotential neuroglial precursor. The authors describe an intraventricular neurocytoma in an 11-year-old boy that showed anaplastic features with widespread necrosis and mitoses, as well as extensive calcification and foci that exhibited marked neuronal differentiation with clusters of ganglion cells. Immunohistochemical examination showed prominent synaptophysin and neurofilament positivity and focal glial fibrillary acidic protein positivity. Electron microscopy revealed abundant neuritic processes with microtubules and dense core granules as well as mature ganglion cells. Flow cytometry studies revealed increased S (7.8%) and G2M (9.7%) phase components. Molecular and cytogenetic studies were undertaken to assess whether there were similarities to two other tumor types that exhibit neuronal differentiation, the neuroblastoma and medulloblastoma. Polymerase chain reaction and fluorescence in situ hybridization (FISH) analysis revealed no evidence of amplification of the MYCN oncogene or chromosome 1p deletion, which are common in neuroblastomas. Chromosomal analysis by G banding revealed a complex karyotype, with counts in the near-diploidy range (45–48). Two chromosomes 1 appeared normal on G banding and FISH analysis, with p58 signals present on the distal p arm of both chromosomes 1; however, three additional copies of distal 1q were present in rearrangements with 4 and 7. Although the histological findings indicate a kinship to the neuroblastoma and medulloblastoma, the central neurocytoma appears to have a different karyotypic profile, although more cases need to be assessed using molecular genetic analysis.

scholarly journals Chickpea genotypes characteristics on resistance to fusarium Fusarium oxysporum f. sp. ciceris

2018 ◽  
Vol 23 ◽  
pp. 166-169
Author(s):  
V. A. Chekalov ◽  
N. E. Volkova
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Markers ◽  
Fusarium Oxysporum ◽  
Molecular Genetic ◽  
Molecular Genetic Analysis ◽  
Chain Reaction ◽  
Resistance Level ◽  
Extraction And Purification ◽  
Polymerase Chain ◽  
Southern Ukraine

Aim. Molecular-genetic analysis of the chickpea genotypes for foc0, foc3, foc4 resistance genes to Fusarium oxysporum f. sp ciceris. Methods. Extraction and purification of DNA, spectrophotometry, polymerase chain reaction, electrophoresis in polyacrylamide gels. Results. 35 chickpea lines and varieties of Ukrainian and foreign breeding characterized according to genotyping on foc0, foc3, foc4 genes of resistance to Fusarium oxysporum f. sp ciceris by the microsatellite markers TA59, TR19 and TR59. Fragments of the expected size for all markers were obtained for samples, for which the resistance level was fixed to certain races. Match between data on the presence of a amplification fragment of a certain size and resistance level among other samples is not found. Conclusions. For 35 chickpea varieties and lines the allele state of foc0, foc3, foc4 genes of resistance to the F. oxysporum f. sp ciceris races 0, 3, 4 is established. The variety ‘Pam’yat’ is recommended as a control of resistance to F. oxysporum f. sp ciceris races 0, 3, 4 in the southern Ukraine conditions. Keywords: chickpea, genes, molecular markers, fusarios, resistance.

Histologic Analysis of Gonadal Tissue in Patients with Ullrich-Turner Syndrome and Derivative Y Chromosomes

2005 ◽  
Vol 8 (2) ◽  
pp. 197-203 ◽  
Author(s):  
L.-C. Horn ◽  
A. Limbach ◽  
W. Hoepffner ◽  
R.B. Tröbs ◽  
E. Keller ◽  
...  
Keyword(s):  
At Risk ◽  
Polymerase Chain Reaction ◽  
Turner Syndrome ◽  
Molecular Genetic ◽  
Germ Cell Tumors ◽  
Molecular Genetic Analysis ◽  
Chain Reaction ◽  
Additional Tool ◽  
Polymerase Chain ◽  
Chromosomal Sequences

To identify patients who had Ullrich-Turner syndrome (UTS) and were at risk for gonadoblastoma or associated germ cell tumors, molecular genetic analysis was carried out to detect Y chromosomal sequences. From peripheral blood samples of 5 patients who had cytogenetically confirmed UTS, genomic DNA was extracted and screened for Y chromosomal sequences by polymerase chain reaction. The morphology of the gonadal tissues was compared with results from polymerase chain reaction. Three phenotypic females showed UTS mosaicism with normal X chromosome accompanied by Y chromosomal material, and 2 patients showed marker chromosomes. Molecular analysis represented loci PABY, SRY, ZFY, TSPY, DYZ3, DYZ1 DXYS, 19Y, DYS-273, DYS-148, DYS218, DYS224, and DYZ1. Three patients showed gonadal tumors (1 with unilateral gonadoblastoma, 1 with unilateral dysgerminoma, and 1 patient had both tumors in 1 gonad). Molecular genetic screening for Y chromosomal sequences may be useful as an additional tool for the identification of patients at risk for a gonadal tumor. Careful, complete processing, including step sectioning, of the gonadectomy specimens to detect small lesions is recommended.

scholarly journals Prevalence of beta thalassemia mutations in population of Gujarat using amplification-refractory mutation system–polymerase chain reaction

2019 ◽  
Vol 6 (8) ◽  
pp. 3294
Author(s):  
Pankaj K. Gadhia ◽  
Salil N. Vaniawala ◽  
Tushar B. Kachhadiya
Keyword(s):  
Polymerase Chain Reaction ◽  
Genetic Analysis ◽  
Prevalence Rate ◽  
Molecular Genetic ◽  
Molecular Genetic Analysis ◽  
Beta Thalassemia ◽  
Amplification Refractory Mutation System ◽  
Chain Reaction ◽  
Mutation System ◽  
Polymerase Chain

Background: Beta thalassemia is the most common genetic disorder in India. Its trait, coinheritance and mutations vary from mild to severe condition, resulting in thalassemia minor (heterozygous), intermediate and major depending upon many factors. The objective of this study was to find out the prevalence rate and the carrier of beta thalassemia in population of Gujarat using molecular genetic analysis of beta thalassemia patients by targeted mutation assay (ARMS-PCR).Methods: A total 105 samples for beta thalassemia were analysed for IVS 1-5 (G→C) and CD 15 (G→A) mutations. These two common mutations of thalassemia in Gujarat were carried out using amplification refractory mutation system–polymerase chain reaction (ARMS-PCR) and gel electrophoresis method.Results: A total 105 samples referred to us for molecular genetic analysis. The occurrence of positive mutations of IVS 1-5 (G→C) and CD 15 (G→A) were found in 48 and 15 samples respectively. The rest were negatives.Conclusions: Present study concludes that the prevalence rate of Beta thalassemia was widespread among the Gujarat population. The identification of IVS 1-5 (G→C) and CD 15 (G→A) mutations was carried out. The analysis revealed that, mutational patterns of IVS 1-5 (G→C) was the most frequent among other mutations in Gujarat region. 

scholarly journals A Comparative Study of 5S rDNA Non-Transcribed Spacers in Elaeagnaceae Species

Plants ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 4
Author(s):  
Oleg S. Alexandrov ◽  
Olga V. Razumova ◽  
Gennady I. Karlov
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Markers ◽  
Dna Markers ◽  
5S Rdna ◽  
Chain Reaction ◽  
Closely Related Species ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Species Specific ◽  
Shepherdia Canadensis

5S rDNA is organized as a cluster of tandemly repeated monomers that consist of the conservative 120 bp coding part and non-transcribed spacers (NTSs) with different lengths and sequences among different species. The polymorphism in the 5S rDNA NTSs of closely related species is interesting for phylogenetic and evolutional investigations, as well as for the development of molecular markers. In this study, the 5S rDNA NTSs were amplified with universal 5S1/5S2 primers in some species of the Elaeagnaceae Adans. family. The polymerase chain reaction (PCR) products of five Elaeagnus species had similar lengths near 310 bp and were different from Shepherdia canadensis (L.) Nutt. and Sh. argentea (Pusch.) Nutt. samples (260 bp and 215 bp, respectively). The PCR products were cloned and sequenced. An analysis of the sequences revealed that intraspecific levels of NTS identity are high (approximately 95–96%) and similar in the Elaeagnus L. species. In Sh. argentea, this level was slightly lower due to the differences in the poly-T region. Moreover, the intergeneric and intervarietal NTS identity levels were studied and compared. Significant differences between species (except E. multiflora Thunb. and E. umbellata Thunb.) and genera were found. Herein, a range of the NTS features is discussed. This study is another step in the investigation of the molecular evolution of Elaeagnaceae and may be useful for the development of species-specific DNA markers in this family.

Automated closed-vessel system for in vitro diagnostics based on polymerase chain reaction

1993 ◽  
Vol 39 (9) ◽  
pp. 1927-1933 ◽  
Author(s):  
J B Findlay ◽  
S M Atwood ◽  
L Bergmeyer ◽  
J Chemelli ◽  
K Christy ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Pcr Amplification ◽  
Automated System ◽  
Closed Vessel ◽  
Chain Reaction ◽  
Panel Testing ◽  
Pcr Product ◽  
Pcr Products ◽  
Polymerase Chain

Abstract An automated system for polymerase chain reaction (PCR) amplification and detection combats false-positive results caused by "PCR product carryover." The system uses a single vessel for both PCR amplification and the subsequent detection of PCR products, eliminating the need to handle PCR products in an open environment and risk product carryover. The sample and PCR reagents are introduced into one compartment within the vessel, and amplification occurs as they are thermally cycled. Other compartments contain the reagents for detection of PCR products. Pressure from a roller provides for sequential delivery of the contents of the compartments to a detection area. The PCR products are biotinylated at their 5' ends during amplification through the use of biotinylated primers. After delivery to the detection area, they are specifically captured by hybridization with immobilized oligonucleotide probes. Subsequent reaction with streptavidin-horseradish peroxidase conjugate forms a complex that catalyzes dye formation from dye precursor. Wash steps minimize nonspecific background. This format is amenable to multiplexing, permitting internal controls, speciation of bacteria, typing of viruses, and panel testing. An HIV assay performed with this system demonstrated 100% sensitivity and 95% specificity for 64 patients' samples relative to a conventional PCR assay based on 32P solution hybridization. Similarly, an automated closed-vessel assay of cytomegalovirus exhibited 97.5% sensitivity and 100% specificity.

scholarly journals Ligation-Mediated Polymerase Chain Reaction Detection of 8-Oxo-7,8-Dihydro-2′-Deoxyguanosine and 5-Hydroxycytosine at the Codon 176 of the p53 Gene of Hepatitis C-Associated Hepatocellular Carcinoma Patients

2020 ◽  
Vol 21 (18) ◽  
pp. 6753
Author(s):  
Andrea Galli ◽  
Armelle Munnia ◽  
Filippo Cellai ◽  
Mirko Tarocchi ◽  
Elisabetta Ceni ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Polymerase Chain Reaction ◽  
Hepatitis C ◽  
Molecular Mechanisms ◽  
Active Oxygen Species ◽  
Dietary Exposure ◽  
P53 Gene ◽  
Chain Reaction ◽  
Specific Sequence ◽  
Polymerase Chain

Molecular mechanisms underlying Hepatitis C virus (HCV)-associated hepatocellular carcinoma (HCC) pathogenesis are still unclear. Therefore, we analyzed the levels of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) and other oxidative lesions at codon 176 of the p53 gene, as well as the generation of 3-(2-deoxy-β-d-erythro-pentafuranosyl)pyrimido[1,2-α]purin-10(3H)-one deoxyguanosine (M1dG), in a cohort of HCV-related HCC patients from Italy. Detection of 8-oxodG and 5-hydroxycytosine (5-OHC) was performed by ligation mediated-polymerase chain reaction assay, whereas the levels of M1dG were measured by chromatography and mass-spectrometry. Results indicated a significant 130% excess of 8-oxodG at –TGC– position of p53 codon 176 in HCV-HCC cases as compared to controls, after correction for age and gender, whereas a not significant increment of 5-OHC at –TGC– position was found. Then, regression models showed an 87% significant excess of M1dG in HCV-HCC cases relative to controls. Our study provides evidence that increased adduct binding does not occur randomly on the sequence of the p53 gene but at specific sequence context in HCV-HCC patients. By-products of lipid peroxidation could also yield a role in HCV-HCC development. Results emphasize the importance of active oxygen species in inducing nucleotide lesions at a p53 mutational hotspot in HCV-HCC patients living in geographical areas without dietary exposure to aflatoxin B1.

Sign in / Sign up

Export Citation Format

Share Document