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At least 703 records · Page 39Linked to original sources
PubMed1994

A common hot spot for somatic and germline mutations in rat mitochondrial DNA: analysis by fluorescence-based SSCP.

To estimate the degree of sequence heterogeneity of mtDNA, we have developed an efficient system for mutant detection using fluorescence-based single strand conformational polymorphism (F-SSCP). The F-SSCP analysis 200 clones from Wistar rat and sequencing of 2 clones from each 40 inbred Wistar rats at the age of 7 weeks detected no sequence differences, suggesting that the mtDNA sequence heterogeneity is low in young rat hearts. One of the 387 clones isolated from a Donryu rat showed a different mobility in the F-SSCP from other clones. Sequencing of the mutant clone revealed double contiguous TT-->CC transitions at nucleotide positions 15443 and 15444, compared with the normal Donryu clone. The nucleotide at 15444 was different between the Wistar and Donryu rats, indicating that this is the site of germline mutation. These results suggest that the somatic and germline mutations can occur at a common hot spot.

Animals↗
PubMed1996

Primary myiasis in dog caused by Phaenicia eximia (Diptera:Calliphoridae) and preliminary mitochondrial DNA analysis of the species in Brazil.

A case of primary myiasis in a dog caused by Phaenicia eximia (Robineau-Desvoidy) in Brazil is presented. A young and healthy female dog, Canis familiaris, approximately 10 d old and still under maternal care, was found to have several eggs and 1st instars larvae in its abdomen and urogenital regions. Samples were collected in Campinas, São Paulo, and transferred to the laboratory for rearing and identification. A comparative analysis of the mitochondrial DNA (mtDNA) with 12 restriction enzymes in 2 sampled populations of P. eximia collected in different hosts (live dog and bovine carcass) and in the same locality revealed that 4, EcoRI, EcoRV, HaeIII, and MspI were suitable for detecting mtDNA markers in the 2 populations.

Animals↗
PubMedDate not supplied

DNA analysis of the fragile X syndrome in an at risk pediatric population in croatia: simple clinical preselection criteria can considerably improve the cost-effectiveness of fragile X screening studies.

Advances in understanding the molecular basis of the fragile X syndrome, the most common cause of inherited mental retardation, have elicited new prospects for population-based studies identifying affected individuals and fragile X families, thus contributing in prevention of the disease. In comparison with numerous fragile X screening studies where unselected groups of individuals with mental retardation, developmental delay, learning disability or autistic-like behaviour had been observed, we performed fragile X analysis on clinically preselected individuals. The group we studied consisted of 108 children with mental retardation of unknown cause or positive family history who had at least one physical and/or behavioural characteristic often observed among fragile X individuals. A relative high frequency of the fragile X positive cases (13% overall, 17.3% in males) was detected, suggesting that simple preselection criteria can considerably increase the proportion of fragile X-positive cases, and therefore, improve the cost-effectiveness of fragile X testing. Retrospective clinical analysis using a simplified six-item fragile X checklist confirmed that scoring criteria can be used to additionally preselect individuals at risk. Our results also indicate that this syndrome is underdiagnosed in Croatia and that a further effort must be made to detect unrecognised cases.

Child↗
PubMed1989

[Prediction of radiosensitivity by DNA analysis].

The aim of the study is to investigate the relationship between DNA histogram and radiosensitivity on tumor specimens taken directly from patients. Flow cytometric DNA content analysis with PI (Propidium iodide) was performed on specimens of cervical carcinoma. Radiosensitivities of respective tumors were determined on biopsy specimens that were taken 3 weeks after the start of radiotherapy. Five parameters (over 4C, %G2M, %S, PI, DI) were calculated from DNA Histogram. Among these parameters over 4C, %G2M and PI correlated well to radiosensitivity.

Adult↗
PubMed1995

[Sisters with early onset hereditary dentatorubral-pallidoluysian atrophy of childhood--DNA analysis and clinicopathological findings].

Two sisters were presented, 16 years old and 12 years old, who showed similar clinical courses. They had had mental retardation since early childhood, and then ataxia began. They suffered from astatic and tonic seizures from early school age, which gradually evolved to intractable epilepsies. Spasticity progressed, and they deteriorated both physically and mentally. They revealed photo-sensitivity; convulsions were induced by the flickering of light. They were attacked by myoclonic seizures as well as choreoathetosis, and became bedridden by the latter part of the elementary school age. There were no fruitful results of any kind from the laboratory examinations for metabolic disorders. EEG showed that the epileptic seizure discharges were induced by photic stimulation; there were frequent 3-4 Hz diffuse spike-and-wave short bursts during waking and sleep periods. MRI findings of the elder sister at the age of 16 revealed remarkable diffuse brain atrophy. Gene analysis showed abnormally enlarged DNA fragments localized on the short arm of chromosome 12. This meant expanded CAG trinucleotide repeats. The younger sister died at the age of 12 years. Autopsy findings revealed degeneration of both dentatorubral and pallidoluysian pathways. There were especially remarkable gliosis and neuronal cell loss in the outer segment of globus pallidus, and moderate neuronal cell loss and typical grumose degeneration in the dentate nucleus. The diagnosis of juvenile-type hereditary dentatorubral-pallidoluysian atrophy was compatible with the pathologic findings. This diagnosis will be made possible before death through the understanding of the clinical symptoms and molecular genetics.

Adolescent↗
PubMedDate not supplied

Direct DNA analysis of fragile X syndrome in Spanish pedigrees.

Eleven complete Spanish pedigrees with fragile X syndrome were analysed by Southern blotting with the DNA probe StB12.3 previously isolated and described by Oberlé et al. [1991]. This probe allowed the direct detection of affected males and carrier females and was able to distinguish between normal males and normal transmitting males (NTMs). One hundred and twenty three individuals were analyzed, 115 from the pedigrees and 8 from the general population. Five mosaic cases were found (4 males and one female) showing both the premutation and the full mutation. One half of the females with the full mutation were mentally retarded but no female with mental retardation carried the premutated pattern, suggesting that the absence of the full mutation in females is a very good criterion for pre-or postnatal diagnosis of normal mental status.

Cytogenetics↗
PubMed2001

Optimization of random amplification of polymorphic DNA analysis for molecular subtyping of Escherichia coli O157.

Random amplification of polymorphic DNA (RAPD) analysis using the polymerase chain reaction has proved to be a useful technique in the epidemiological investigation of micro-organisms but may suffer from a lack of reproducibility in poorly optimized protocols. In this study a method of obtaining reproducible genomic fingerprints using RAPD analysis of Escherichia coli O157 is described. By systematic optimization of reaction conditions and selection of suitable primers, reproducible and discriminatory profiles could be obtained from all E. coli O157 strains tested. In addition, two other methods of obtaining reproducible profiles from E. coli O157 strains without the need to purify genomic DNA are described.

Buffers↗
PubMed1994

[Flow cytometric DNA analysis and chemosensitivity in squamous cell carcinoma of the head and neck].

Cellular DNA content and succinate dehydrogenase activity of 92 human head and neck (34 laryngeal, 24 pharyngeal, 21 oral cavity, 13 maxillary) squamous cell carcinomas were examined, and DNA ploidy status and chemosensitivity were analyzed and compared. DNA aneuploidy was observed in 54 tumors (58.7%). The aneuploid pattern was most common in tumors of the maxillary sinus (84%), and least common in tumors of the larynx (41.3%). Histologically, aneuploidy was detected in 71.4% of poorly-differentiated, 63.8% of moderately-differentiated and 37.5% of well-differentiated squamous cell carcinomas. There was a statistically significant difference between the survival rates of patients with diploid and aneuploid patterns. Chemosensitivity was determined by exposing fresh tumor material to five antitumor drugs: adriamycin (ADM), cisplatin (CDDP), carboquone (CQ), 5-fluorouracil (5-FU) and mitomycin C (MMC). The average decrease in succinate dehydrogenase (SD) activity was 49.8% with ADM, 33.6% with CDDP, 39.9% with CQ, 68.4% with 5-FU and 45.5% with MMC. Histologically, poorly-differentiated squamous cell carcinomas were most sensitive to these five antitumor drugs. We also compared average SD activity in tumors from different organs and found that pharyngeal tumors tend to be most sensitive to these drugs, except for MMC. The chemosensitivity of a tumor with DNA diploidy tended to be higher among well- and moderately-differentiated squamous cell carcinomas. In contrast, tumors with DNA aneuploidy tended to have higher chemosensitivity in the poorly-differentiated type. The results of this study indicate that simultaneous analysis of DNA ploidy and chemosensitivity will be helpful in understanding the characteristics of tumors as well as in predicting the most effective chemotherapy agents for head and neck cancer patients.

Adult↗
PubMed2000

Sex determination based on fecal DNA analysis of the amelogenin gene in sika deer (Cervus nippon).

A sex determination method using DNA extracted from feces has been developed for sika deer (Cervus nippon). We determined a partial sequence of the amelogenin gene of sika deer, which exists on both X and Y chromosomes with a deletion region on the Y chromosome. Based on the sexually dimorphic sequences, we designed a pair of primers which could amplify DNA fragments the lengths of which are different between males and females. PCR products were detected in 34 out of 37 fecal samples collected from captured deer and the sexes estimated by the present method were perfectly matched with the actual sexes.

Amelogenin↗
PubMed1992

Entamoeba histolytica extrachromosomal circular ribosomal DNA: analysis of clonal variation in a hypervariable region.

The ribosomal RNA genes of the protozoan parasite Entamoeba histolytica are highly repeated and display restriction fragment length polymorphism. Using a set of four DNA probes spanning the coding region and part of the flanking region of the E. histolytica ribosomal RNA genes, an analysis of the DNA bands generated by EcoRI digestion of Entamoeba DNA is presented. This analysis included five strains of E. histolytica, four strains of E. moshkovskii, and one strain each of E. invadens and E. terrapinae. No common bands were observed between E. histolytica and the other Entamoeba. Within E. histolytica, two bands were conserved in all strains while the others were polymorphic. Detailed analysis of DNA from independently isolated clones of the strain HM-1:IMSS of E. histolytica showed two bands to be highly polymorphic. Of these, the 4.4-kb band of clone 6 was further analyzed. Polymorphism in this band could even be demonstrated in cells of the same clone. Restriction enzyme analysis of this DNA band from two clones of HM-1:IMSS showed that the polymorphism may be due to variable numbers of DraI repeat units present in this DNA stretch.

Animals↗
PubMed2005

Forensic mitochondrial DNA analysis of 691 casework hairs.

A five year retrospective review of mitochondrial DNA (mtDNA) analysis on 691 casework hairs was carried out. A full or partial mtDNA profile was obtained for > 92% of hairs. With increasing age of the hair, the likelihood of obtaining a full profile decreased, although "mini-primer sets" could often be used to capture a partial profile. With increasing color and diameter of the hair, the likelihood of obtaining a profile increased. Full or partial profiles were obtained on more than 80% of 114 hairs < or = 1.0 cm. Mixtures were observed in 8.7% of hairs tested; mixtures increased with the age of the hair and were presumed to be due to exterior surface contamination that could not be sufficiently cleaned prior to extraction, since the overall level of laboratory contamination was low. The frequency of sequence heteroplasmy was 11.4%, and both hot-spot and novel sites were observed. In about one-third of these observations, another sample in the case showed either the same heteroplasmic site or a nucleotide substitution at that site.

DNA Fingerprinting↗
PubMed2003

Automated high-throughput probe production for DNA microarray analysis.

DNA microarrays have become an established tool for gene expression profiling. Construction of these microarrays using immobilized cDNAs is a common experimental strategy. However, this is extremely laborious, requiring the preparation of hundreds or thousands of cDNA probes. To minimize this initial bottleneck, we developed a comprehensive high-throughput robotic system to prepare DNA probes suitable for microarray analysis with minimal user intervention. We describe an automated system using the MultiPROBE Nucleic Acid Purification Workstation to provide the liquid handling and other functions needed to optimize this process. We were able to carry out fully automated plasmid cDNA isolation, PCR assay setup, and PCR purification and also to direct the characterization and tracking of DNA probes during processing. Protocols began with the initial preparation of a plasmid DNA archive of bacterial stocks in parallel 96-well plates (192 samples/run) and continued through to the dilution and reformatting of chip-ready DNA probes in 384-well format. These and other probe production procedures and additional instrument systems were used to process fully a set of mouse cDNA clones that were then validated by differential gene expression analysis.

Animals↗
PubMed1981

Comparative single-cell and flow DNA analysis of bone sarcoma.

Five chondrosarcomas and seven osteosarcomas were analyzed by single-cell DNA cytophotometry, in both imprint preparations and tissue sections, and by means of flow DNA cytophotometry. A comparative analysis of the DNA distribution curves of each tumor, as determined by these three methods, showed a good correlation with respect to the modal DNA values. In addition, there was a strong correlation between the proportion of tumor cells with an increased DNA value (hyperploid cells) in imprint preparations and in tissue sections of each tumor as determined by single-cell DNA cytophotometry. Flow cytophotometry appears to be the method of choice for the determination of the aneuploid modal DNA value of a tumor cell population, while single-cell cytophotometry of identified cells seems more appropriate for the determination of the proportion of hyperploid cells in a tumor cell population. In this respect, single-cell and flow DNA cytophotometry may be considered complementary. For the demonstration of the existence of aneuploidy in a tumor cell population, however, the three investigated methods seem to be equivalent. Previous studies using single-cell cytophotometry have shown that DNA determinations in chondrosarcoma as well as in other tumors provide significant prognostic information. For routine clinical work, however, as an adjunct to the conventional histopathologic malignancy grading, single-cell cytophotometry is a rather time consuming procedure, while flow cytophotometry offers a rapid and convenient means of determining the nuclear DNA content. The present study clearly showed that flow DNA cytophotometry may be applied to sarcoma of bone.

Bone Neoplasms↗
PubMed1986

Clinical application of DNA analysis in a family with OTC deficiency.

We describe the clinical application of DNA restriction fragment analysis to the genetic evaluation of a family with a child deficient in ornithine transcarbamylase (OTC). The results of protein loading studies and the interpretation of the DNA haplotype profiles for the human OTC gene are reported. DNA restriction fragment analysis may be a reliable technique for the prenatal diagnosis of OTC deficiency and identification of obligate carriers of this gene.

Amino Acid Metabolism, Inborn Errors↗
PubMed1995

Detection of chromosome aneuploidy in breast lesions with fluorescence in situ hybridization: comparison of whole nuclei to thin tissue sections and correlation with flow cytometric DNA analysis.

We compared flow-cytometric DNA histogram pattern to counts of 4 fluorescent-labelled centromeric probes (chromosomes 1, 7, 8, and 17) in whole nuclei (WN) and in nuclei from the corresponding formalin-fixed deparaffinized thin tissue section (TS) in 25 breast lesions (9 invasive carcinomas, 1 duct carcinoma-in-situ, 5 fibroadenomas, 10 fibrocystic change). In benign lesions, signal gains (i.e., trisomic nuclei) were never observed in greater than 10% of nuclei from either WN or TS preparations. Loss of signal in benign breast lesions, however, varied considerably (0-43%) between individual case and between chromosome probes. The mean incidence of signal loss in WN of benign lesions ranged from 8.9% (chromosome 7) to 14.4% (chromosome 1) of nuclei. These signal loss frequencies exceeded those of benign lymphoid control cells. In three benign lesions, signal loss in WN (with one probe) was observed in at least 25% of nuclei. Signal losses in benign TS, on average, were 50-150% greater than in matched WN preparations (chromosome 1-21.7%, chromosome 7-21.5%). Malignant lesions generally, but not always, displayed fewer monosomic nuclei and more trisomic nuclei in WN compared to TS, compatible with a slicing (i.e., nuclear truncation) artifact. Signal counts in carcinomas correlated well with flow cytometric DNA index; however, they were also characterized by evidence of genetic instability, manifest as signal gains in a subset of nuclei (10-25%) with individual probes in diploid range cases, as well as intratumoral heterogeneity, reflected as discrepancies in probe counts between WN and TS samples. We conclude that signal losses with centromeric probes are largely, but not entirely, explained by nuclear slicing.(ABSTRACT TRUNCATED AT 250 WORDS)

Aneuploidy↗
PubMed1994

Hb Lepore Baltimore (delta 50Ser beta 86Ala) identified by DNA analysis in a Brazilian family.

Genomic DNA from two Brazilian hemoglobin (Hb) Lepore heterozygotes of Italian ancestry have been studied in order to identify the Hb Lepore type and to sequence the breakpoint region. The two genes were sequenced after PCR amplification and had the delta globin sequence up to exon 2 codon 68 while the first specific base for the beta globin gene was at codon 86 of the second exon; between the two ends, they had 51 base pairs in common with the delta and beta globin genes. These data indicate that the mutation was of the Hb Lepore Baltimore type. The Lepore chromosome haplotype was different from that previously described in members of a Spanish family with Hb Lepore Baltimore. These data suggest that independent mutations have given rise to Hb Lepore Baltimore in different regions of the world.

Anemia↗
PubMed1991

[Flow cytometric DNA analysis of uterine endometrial carcinoma].

The DNA content in individual cells from 40 cases of histopathologically normal endometria, 15 of endometrial hyperplasia and 68 of endometrial adenocarcinoma was measured by flow cytometry. An aneuploid cell population was found in 50% of malignant endometria, but in the remaining endometrial carcinomas, the flow cytometrical findings showed no difference from those of benign tissue. Aneuploidy was more common (77.8%) in poorly differentiated tumors than in highly differentiated tumors (35.5%). Two and more aneuploid cell populations were found in 8 cases of 34. The DNA indices of aneuploid tumors were classified into 3 groups: hyperdiploidy, low hyperdiploidy (DNA index range: 1.04-1.2) and high hyperdiploidy (DNA index range: More than 1.2). The proportion of tumors with a high DNA index tended to increase as tumors became less differentiated. In normal endometria the fraction of cells with DNA content corresponding to the s-phase (s-fraction) was 8 +/- 3% on average in the proliferative phase. In well differentiated diploid tumors the s-fraction was 12 +/- 6%, but in moderately and poorly differentiated tumors it was higher (16.0% and 19.0%).

Adenocarcinoma↗