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Results for “Nested Genes”

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At least 865 records · Page 48Linked to original sources

PubMed1994

Gene expression of interstitial collagenase (matrix metalloproteinase 1) in gastrointestinal tract cancers.

Collagenolytic proteinases play an important pathological role in the invasion and metastasis of cancer cells. However, little is known about the role of interstitial collagenase (matrix metalloproteinase-1; MMP-1) in this process. To investigate the expression of the MMP-1 gene in cancer tissues, an in situ hybridization study was carried out in gastrointestinal tract cancers (one esophageal cancer, five gastric cancers, and four colorectal cancers), using a 35S-labeled MMP-1 cDNA probe. The MMP-1 gene was expressed in the stromal cells of fibrous tissue around cancer nests, especially at the margin of invasion and/or within the cancer nest; however, no definite expression within cancer cells was observed. Expression of the MMP-1 gene in the stromal cells was more common in well differentiated gastric adenocarcinoma than in poorly differentiated adenocarcinomas. These findings indicate that expression of the MMP-1 gene is greater in stromal cells that are closely associated with cancer cells, suggesting a pathophysiological role of MMP-1 in the invasion and metastasis of cancer cells.

Adenocarcinoma↗

PubMed2003

Short communication: point mutations in the dihydrofolate reductase and dihydropteroate synthase genes of Plasmodium falciparum isolates from Colombia.

Point mutations in the dihydrofolate reductase (DHFR) and dihydropteroate synthase (DHPS) genes of Plasmodium falciparum can lead to an increasing resistance of P. falciparum to pyrimethamine/sulphadoxine. We examined the prevalence of these mutations in 36 samples from Colombia. Analysed by polymerase chain reaction (PCR) for infection with P. falciparum, 25 (69%) tested positive. These positive isolates were tested further for point mutations in the genes of DHFR (codons 16, 51, 59, 108 and 164) and DHPS (codons 436, 437, 540, 581 and 613) by nested PCR and following mutation-specific restriction enzyme digestion. Gene mutations occurred in both the DHFR and DHPS gene of the Colombian isolates, suggesting that resistance to antifolate drugs exists or may develop soon in Colombia.

Adult↗

PubMed1992

Rapid method for detecting monoclonality in B cell lymphoma in lymph node aspirates using the polymerase chain reaction.

AIMS: To use the polymerase chain reaction to detect monoclonality at the immunoglobulin heavy chain gene locus in cells derived from lymph node aspirates. METHODS: A nested two-stage polymerase chain reaction (PCR) for the VDJ region of the immunoglobulin heavy chain gene was used to detect monoclonality. The total number of cells available for diagnosis by PCR in lymph node aspirates was between 10(4) and 10(5). RESULTS: A monoclonal band was detected in 21 of 25 malignant B-lymphomas. The other four specimens gave polyclonal bands. Specimens from reactive lymph nodes produced polyclonal bands in 14 cases, no product in two cases, and one specimen gave two monoclonal bands. Polyclonal bands were obtained for three Hodgkin's lymphoma samples and five metastatic carcinomas. Four metastatic carcinoma samples gave no amplification. CONCLUSIONS: Detection of monoclonality in a cell population is strongly suggestive of malignant disease. The simple PCR method presented here should complement conventional cytological and immunological methods for diagnosis of malignancy by lymph node aspirates.

Base Sequence↗

PubMed2005

An antisense RNA plays a central role in the replication control of a repC plasmid.

The widespread replicons of repABC and repC families from alpha-proteobacteria share high similarity in their replication initiator proteins (RepC). Here we describe the minimal region required for stable replication of a member of the repC family, the low copy-number plasmid pRmeGR4a from Sinorizobium meliloti GR4. This region contains only two genes: one encoding the initiator protein RepC (46.8 kDa) and other, an antisense RNA (67 nt). Mapping of transcriptional start sites and promoter regions of both genes showed that the antisense RNA is nested within the repC mRNA leader. The constitutively expressed countertranscribed RNA (ctRNA) forms a single stem-loop structure that acts as an intrinsic rho-independent terminator. The ctRNA is a strong trans-incompatibility factor and negative regulator of repC expression. Based on structural and functional similarities between members of the repC and repABC families we propose a model of their evolutionary relationship.

Amino Acid Sequence↗

PubMed2004

Identification of gammadeltaT lymphocytes in human periapical lesions.

Endodontic (root canal) therapy is required when the pulp of a tooth becomes necrotic due to a bacterial infection or trauma. A proportion of patients who receive endodontic therapy subsequently have periapical (around the tooth root) lesions detected by radiolucency. Currently, there are no means to identify susceptible patients. Although tissue from periapical lesions has been described as inflammatory, inflammatory cell types and their functions have been poorly characterized. For example, T lymphocytes were identified using pan specific anti-CD3 mAb, which recognizes both alphabeta and gammadeltaT cells. Using the current model of gammadeltaT cells as immunoregulatory cells; gammadeltaT cells can mediate protective or destructive milieus. We postulated that patients who have a periapical lesion, as identified by radiographic bone loss, mount a gammadeltaT cell response. We collected specimens removed by surgery from both periapical lesions and other oral tissues, generated total RNA and performed reverse-transcriptase polymerase chain reaction to identify rearranged delta genes. Results were confirmed with semi-nested polymerase chain reaction. In addition, we demonstrate that these lesions contain a population of CD3+ cells that are alphabetaT cell receptor negative, implying that these cells are gammadeltaT cells. Here we show that 36/37 of periapical lesions and only 2/11 of other lesions contain gammadeltaT cells (P<0.0001). Vdelta2+ T cells were the most common subtype identified (30/36) in these samples. This is the first report in the literature of the presence of gammadeltaT cells in human periapical lesions.

Adult↗

PubMed2006

How corridors reduce indigo bunting nest success.

Corridors are a popular strategy to conserve biodiversity and promote gene flow in fragmented landscapes. Corridor effectiveness has been bolstered by the fact that no empirical field studies have shown negative effects on populations or communities. I tested the hypothesis that corridors increase nest predation in connected habitat fragments relative to unconnected fragments. I evaluated this hypothesis in a large-scale experimental system of open-habitatfragments that varied in shape and connectivity. Corridors increased nest predation rates in connected fragments relative to unconnected fragments with lower edge:area ratios. Nest predation rates were similar between connected and unconnected fragments with higher edge:area ratios. These results suggest that the increase in predator activity is largely attributable to edge effects incurred through the addition of a corridor This is the first field study to demonstrate that corridors can negatively impact animal populations occupying connected fragments.

Animals↗

PubMed2005

Plasminogen activator inhibitor-1 4G/5G polymorphism and risk of stroke: replicated findings in two nested case-control studies based on independent cohorts.

BACKGROUND AND PURPOSE: Impaired fibrinolytic function secondary to elevated plasminogen activator inhibitor-1 (PAI-1) levels has been implicated in ischemic stroke. PAI-1 levels are determined by genetic factors and environmental factors, triglyceride levels in particular. The aim of this study was to investigate the common functional 4/5 guanosine (4G/5G) polymorphism in the promoter region of the PAI-1 gene and the risk of stroke. METHODS: A nested case-control study design was applied, using baseline data for 2 independent cohorts obtained at population-based surveys in northern Sweden. In study A, there were 113, and in study B, there were 275 individuals without major concomitant disease at baseline who later experienced a first-ever stroke. Blood samples obtained at baseline were analyzed for potential risk factors, including the 4G/5G polymorphism of the PAI-1 gene. RESULTS: The 4G allele of the PAI-1 polymorphism was associated with an increased risk of future ischemic stroke in both studies (odds ratio [OR] of 4G homozygosity, 1.87; 95% CI, 1.12 to 3.15 in study A; OR of 4G homozygosity, 1.56; 95% CI, 1.12 to 2.16 in study B). Individuals with the combination of hypertriglyceridemia and 4G homozygosity were at the greatest risk of developing stroke. Multiple logistic regression analysis identified 4G homozygosity, systolic blood pressure, and diabetes as independent predictors of ischemic stroke. CONCLUSIONS: Identical findings in 2 independent studies strongly suggest a true and clinically important association between PAI-1 4G/5G genotype and risk of future ischemic stroke. The observed modification of the genotype effect by triglycerides may be interpreted as a gene-environment interaction.

Adult↗

PubMed1995

RT/PCR detection of SIL-TAL-1 fusion mRNA in Chinese T-cell acute lymphoblastic leukemia (T-ALL).

The TAL-1 gene is located on chromosome 1p32. In about 20% of T-cell acute lymphoblastic leukemias (T-ALL), this gene is disrupted in its 5' portion by a site-specific 100-kg deletion and is fused with the 5' part of the SIL gene, to form SIL-TAL-1 chimeric gene. In this study, we established a "nested" retrotranscriptase/polymerase chain reaction (RT/PCR) technique which allows detection of the SIL-TAL-1 transcriptional expression. A chimeric mRNA was observed in four of 17 T-ALL cases and has been shown to result from the fusion between the exon 1 of SIL and exon 3 of TAL. A sensitivity test showed that this RT/PCR procedure could detect one leukemic cell among 10(6) normal cells. A positive RT/PCR result was obtained in two cases during clinical remission, suggesting the presence of minimal residual disease (MRD). One patient developed clinical relapse 3 months after PCR positivity. Moreover, analysis of the Tald rearrangement by DNA-based PCR in four patients with SIL-TAL-1 fusion revealed the type A (Tald1) rearrangement in all cases. Sequence analysis demonstrated the presence of N region and non-random "P" nucleotide, as well as base deletions at the genomic SIL-TAL-1 joining site. These data indicate that detection of TAL-1 gene abnormality is important for diagnosis and monitoring of MRD in a subset of T-ALL.

Adolescent↗

PubMed1996

AZT-related mutation Lys70Arg in reverse transcriptase of human immunodeficiency virus type 1 confers decrease in susceptibility to ddATP in in vitro RT inhibition assay.

The genetic basis for didanosine (ddl) resistance in human immunodeficiency virus (HIV-1) has previously been shown to be commonly associated with a Leu to Val change at codon 74 in the HIV-1 RT gene. In this study sequential viral isolates were analyzed from five patients with prior zidovudine (AZT) use who received 6 to 16 months of ddl therapy. Following ddl therapy, viral isolates exhibited an increased AZT susceptibility and decreased ddl susceptibility. Sequence and nested PCR analysis of the HIV-1 RT gene revealed that two viral isolates contained the Leu to Val change at codon 74, and three other isolates with reduced susceptibility to ddl each contained changes at codons 65, 70, and 72. Site-directed mutagenesis was employed to insert specific mutations in RT gene of proviral clone pNL4-3. Analysis of virion-associated reverse transcriptase activity indicated that the Lys70Arg mutation resulted in an enzyme with 2- to 4-fold decreased susceptibility to ddATP. Statistical analysis of the inhibitory concentration for RT activity between pNL4-3 and mutant Lys70Arg viruses obtained in three independent RT inhibition assays was significant (P = 0.05) by student t test paired analysis. Drug susceptibility assays on the virus with Lys70Arg mutation showed a marginal decrease in susceptibility to ddl (1.5- to 2-fold) and about 4- to 6-fold decrease in susceptibility to AZT. Mutations Lys65Glu and Arg72Ser resulted in an impaired RT with greatly diminished functional RT activity. The AZT-associated Lys70Arg mutation results in an RT enzyme with decreased susceptibility to ddATP.

Base Sequence↗

PubMed2002

Evidence that superficial basal cell carcinoma is monoclonal from analysis of the Ptch1 gene locus.

BACKGROUND: Superficial basal cell carcinoma (BCC) arises as an apparently multifocal proliferation of tumour nests attached to the epidermis, which is at odds with a monoclonal origin. Computer-assisted reconstruction shows that these nests join in three dimensions, but it remains unknown whether this tumour is monoclonal. An early event in BCC formation, loss of heterozygosity (LOH) at the Patched 1 (Ptch1) locus, can be used as a tool to address whether this tumour is monoclonal. OBJECTIVES: To determine whether superficial BCC is monoclonal by analysing individually microdissected superficial BCC nests and looking for the same pattern of LOH in each. METHODS: Six cases of superficial BCC were analysed for LOH at the Ptch1 gene locus using the D9S287 microsatellite marker. Identical allelic patterns were sought in each nest from a given tumour. These patterns were no allelic loss, loss of the shorter allele or loss of the longer allele, each with a respective probability of occurrence, as estimated from published findings. RESULTS: All cases were informative. Four cases showed no LOH in each nest and two showed loss of the same allele. If these nests arose independently, then the probability of this result was between 4 x 10-11 and 2 x 10-14. The lack of LOH, seen in four cases, could be due to monoclonal expansion of a cell retaining both D9S287 alleles, or due to a polyclonal proliferation. Therefore, a separate analysis excluding these cases was done, giving a probability of between 2.2 x 10-4 and 1.0 x 10-7. CONCLUSIONS: These probabilities were so extreme that it was unlikely that the nests arose independently, thus providing the first molecular evidence that superficial BCC is monoclonal.

Basal Cell Carcinoma↗

PubMed2005

[Genotyping of ampicillin-resistant Haemophilus influenzae].

OBJECTIVE: Haemophilus (H.) influenzae is a gram-negative bacillus that is a common commensal organism of the human upper respiratory tract and an important cause of human diseases such as pneumonia, meningitis, septicemia, epiglottitis and cellulitis. Strains of H. influenzae are classified according to their capsular polysaccharide. There are six serotypes, designated as a through f. In addition, there are nonencapsulated strains. Although the type of infectious diseases caused by H. influenzae has changed considerably in recent years because of the widespread and routine immunization of children against type b H. influenzae (Hib), Hib remains an important pathogen. Ampicillin is the drug of choice for treating many infections caused by H. influenzae, but its usefulness has been compromised by the increasing prevalence of ampicillin-resistant strains. The continued monitoring of resistant strains by using genotyping methods may provide insights into the epidemiology of transmission. A molecular epidemiological study of ampicillin-resistant H. influenzae derived from nasopharyngeal swabs specimens of children less than 5 years of age with respiratory tract infection were investigated in this study. METHODS: A total of 899 isolates were collected from Beijing, Shanghai, and Guangzhou during 2000-2003. Susceptibility to ampicillin was determined by using E-test. Ampicillin-resistant H. influenzae strains were selected according to National Committee for Clinical Laboratory Standards (NCCLS) 2002 breakpoints. Nested PCR method with primers specific for bexA gene and b capsulate type-specific gene was established. Genotyping by pulsed-field gel electrophoresis (PFGE) and multiplex PCR assay was performed for all ampicillin-resistant H. influenzae strains. RESULTS: Seventy-four ampicillin-resistant H. influenzae strains were obtained. Two strains were positive by nested PCR, characterized as b genotype. The incidence of Hib in ampicillin-resistant H. influenzae strains was 2.7%; 38 genotypes were detected by PFGE. Detection of five types strains of clonal dissemination by PFGE accounted for 55.4% in all ampicillin-resistant H. influenzae strains. Among them eighteen H. influenzae strains belonged to one type, accounted for 24.3% in all ampicillin-resistant H. influenzae strains. Thirty one genotypes were identified by multiplex PCR assay for ampicillin-resistant H. influenzae. The identity ratio of PFGE and multiplex PCR was 63.5%. CONCLUSION: In Beijing, Shanghai and Guangzhou areas 55.4% of ampicillin-resistant H. influenzae strains had clonal dissemination during the 4 years.

Ampicillin Resistance↗

PubMed2005

Anaplasma platys: an improved PCR for its detection in dogs.

This study compares two PCR assays for the detection of Anaplasma platys in dog blood using primers based on the A. platys 16S rRNA gene. The first approach utilized a "standard" PCR protocol composed of a "single-step" direct amplification using an Ehrlichia genus-specific primer set. The second assay being a "nested" PCR screen that first involved a universal bacterial primer set that amplified the majority of the 16S rRNA gene, followed by the nested round of PCR using an A. platys-specific primer set. Of the 22 dogs sampled, 10 were found to contain A. platys DNA using both protocols, and an additional two dogs were found positive using the nested technique. An extract of A. platys positive genomic DNA was serially diluted and comparison of sensitivities determined between the nested PCR, and a direct assay using A. platys-specific primers. The nested protocol demonstrated an increased sensitivity by at least 2 orders of magnitude when compared to the direct assay alone. Our results indicated that the nested PCR assay with its increased sensitivity would be useful for experimental research investigations as well as offer the potential for use as a routine test in diagnostic pathology.

Anaplasma↗

PubMed1998

Association between glutathione S-transferase M1, P1, and T1 genetic polymorphisms and development of breast cancer.

BACKGROUND: Glutathione S-transferases (GSTs) are encoded by a superfamily of genes and play a role in the detoxification of potential carcinogens. In a nested case-control study, we investigated associations between genetic variability in specific GST genes (GSTM1, GSTT1, and GSTP1) and susceptibility to breast cancer. METHODS: In 1989, a total of 32 898 individuals donated blood samples to a research specimen bank established in Washington County, MD. Genotypes of blood specimen DNA were determined for 110 of 115 women with incident cases of breast cancer diagnosed during the period from 1990 through 1995 and up to 113 of 115 control subjects. Associations between specific genotypes and the development of breast cancer were examined by use of logistic regression to calculate odds ratios (ORs) and 95% confidence intervals (CIs). RESULTS: The GSTM1 homozygous null genotype was associated with an increased risk of developing breast cancer (OR = 2.10; 95% CI = 1.22-3.64), principally due to an association with postmenopausal breast cancer (OR = 2.50; 95% CI = 1.34-4.65). For GSTP1, the data were suggestive of a trend of increasing risk with higher numbers of codon 105 valine alleles (compared with isoleucine alleles); a 1.97-fold increased risk of breast cancer (95% CI = 0.77-5.02) was associated with valine/valine homozygosity. The risk of breast cancer associated with the GSTT1 homozygous null genotype was 1.50 (95 % CI = 0.76-2.95). The risk of breast cancer increased as the number of putative high-risk genotypes increased (P for trend <.001) (OR = 3.77; 95% CI = 1.10-12.88 for a combined genotype of GSTM1 null, GSTT1 null, and either GSTP1 valine heterozygosity or GSTP1 valine homozygosity). CONCLUSIONS: Our findings suggest that genetic variability in members of the GST gene family may be associated with an increased susceptibility to breast cancer.

Breast Neoplasms↗

PubMed1996

The structural gene for rusticyanin from Thiobacillus ferrooxidans: cloning and sequencing of the rusticyanin gene.

The rusticyanin gene from the acidophilic chemolithotroph Thiobacillus ferrooxidans has been cloned and sequenced. A central portion of the gene was identified by PCR reactions utilising primers optimised for codon bias followed by nested PCR with degenerate primers. The 5' and 3' ends of the rusticyanin gene were then cloned using degenerate primers to each end and anchor sequences to the known internal sequence. The entire gene was amplified using Tli DNA polymerase and specific primers to the 5' and 3' ends and the sequence confirmed after cloning into Bluescript and transformation of XL-1 Blue Escherichia coli.

Amino Acid Sequence↗

PubMed2006

Drug transporter and metabolizing enzyme gene variants and nonnucleoside reverse-transcriptase inhibitor hepatotoxicity.

This nested case-control study examined relationships between MDR1, CYP2B6, and CYP3A4 variants and hepatotoxicity during antiretroviral therapy with either efavirenz- or nevirapine-containing regimens. Decreased risk of hepatotoxicity was associated with MDR1 3435C-->T (odds ratio, 0.254; P=.021). An interaction between MDR1 and hepatitis B surface antigen status predicted risk with 82% accuracy (P<.001).

Adult↗

PubMed1998

A nested PCR for detection of North American isolates of bluetongue virus based on NS1 genome sequence analysis of BTV-17.

A nested polymerase chain reaction (PCR)-based assay, for detection of bluetongue virus (BTV) ribonucleic acid in cell culture and tissue samples, was developed. Two pairs of oligonucleotide primers (BTV1 and BTV4 and BTV2 and BTV3), selected from non-structural protein 1 (NS1) gene of BTV-17, were used for the nested PCR in two amplification steps. First a 826-bp product was amplified using an outer primer pair BTV1 and BTV4. The second amplification, using nested or internal primer pair BTV2 and BTV3, produced a 517-bp PCR product. RNA from North American prototype serotypes 2, 10, 11, 13 and 17, propagated in cell cultures, were detected by this nested PCR-based assay. The nested primers BTV2 and BTV3 increased the sensitivity of the BTV PCR assay, and as little as 0.1 fg of BTV RNA (equivalent to 5 viral particles) could be detected. Amplification products were not detected when the PCR-based assay was applied to RNA from a closely related orbivirus, epizootic hemorrhagic disease virus (EHDV) prototype serotypes 1 and 2; total nucleic acid extracts from uninfected BHK-21 cells; or whole blood from calves and deer that were BTV-seronegative and virus isolation negative. Application of this nested BTV PCR-based assay to clinical samples resulted in detection of BTV RNA from a variety of tissues collected from calves and deer with natural and experimental BTV infections. The described BTV PCR-based assay provides a valuable tool to study the epidemiology of BTV infection in susceptible wild ruminants and domestic livestock.

Animals↗

PubMed1995

Multiplex polymerase chain reaction to detect toxigenic Vibrio cholerae and to biotype Vibrio cholerae O1.

A multiplex polymerase chain reaction (PCR) was developed to identify cholera toxin-producing Vibrio cholerae and to biotype V. cholerae O1. Enterotoxin-producing V. cholerae strains were identified with a primer pair that amplified a fragment of the ctxA2-B gene. Vibrio cholerae O1 strains were simultaneously differentiated into biotypes with three primers specified for the hylA gene in the same reaction. The hlyA amplicon in the multiplex PCR serves as an internal control when testing toxin-producing strains, as hlyA gene sequences exist in all tested V. cholerae strains. Enrichment of V. cholerae present on oysters for 6 h in alkaline peptone water before detection by a nested PCR with internal primers for ctxA2-B gene yielded a detection limit lower than 3 colony-forming units (cfu) per gram of food.

Animals↗

PubMed2004

Gene by smoking interaction: evidence for effects on low-density lipoprotein size and plasma levels of triglyceride and high-density lipoprotein cholesterol.

We seek to determine whether significant gene x smoking interaction effects exist on plasma triglyceride (TG) levels, HDL cholesterol (HDL-C) level, and median LDL particle diameter (LDL-MPD) in Mexican American families enrolled in the San Antonio Family Heart Study. The sample consisted of 1,392 individuals distributed in 42 extended pedigrees, ranging in age from 16 years to 92 years. Separate quantitative genetic analyses were carried out for TG and HDL-C level and LDL-MPD using a maximum-likelihood-based variance decomposition approach while simultaneously adjusting for age and sex. Initial heritability estimates demonstrated significant (p < 0.001) additive genetic contributions to all three traits (h2 range 0.50 - 0.54). To test for a gene x smoking interaction, we included in the model additional smoking-status-specific variance terms and a genetic correlation term between smokers and nonsmokers. Comparisons of nested models revealed significant evidence (p < 0.01) for a gene X smoking interaction effect on TG level and LDL-MPD and possible evidence for an effect on HDL-C level. These results indicate that the gene or suite of genes regulating each of these phenotypes is likely the same in smokers and non-smokers but that smoking may alter the expression of genes, particularly those influencing TG level and LDL-MPD.

Adolescent↗