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Cellular and molecular characteristics of an immortalized ataxia-telangiectasia (group AB) cell line.

Ataxia-telangiectasia (A-T) is a multisystem hereditary disease featuring neurodegeneration, immunodeficiency, extreme cancer proneness, chromosomal instability, and radiosensitivity. A-T is found in many ethnic groups, and is genetically heterogeneous: four complementation groups have been identified in A-T so far. Attempts to isolate the A-T gene are based in part on gene transfer experiments, using permanent A-T fibroblast lines, obtained by transformation with SV40. "Immortalization" of A-T primary diploid fibroblasts using SV40 is difficult, possibly because of the chromosomal instability

Improved sensitivity of a modified polymerase chain reaction amplified DNA probe in comparison with serial tissue culture passage for detection of Chlamydia trachomatis in conjunctival specimens from nepal.

A sensitive means for detecting ocular chlamydial infections is needed to accurately define the epidemiology of trachoma. Tissue culture is considered the "gold standard," yet it is less than 50% sensitive for ocular specimens. The purpose of this study was to improve the detection rate of culture by serial passage and thereby provide a more reliable basis for comparing polymerase chain reaction (PCR) amplified and 32P DNA probes and direct fluorescent antibody (DFA) tests with culture. Ocular exams on 1043 individuals were scored for trachoma; 252 (24%) had moderate/severe intensity. A total

Remarkable response of gastric adenocarcinoma with FGFR2-TRIM44 fusion to pemigatinib: a case report.

Fibroblast growth factor receptor 2 (FGFR2) gene fusions are detected in 10%-16% of cholangiocarcinomas but are rarely detected in other solid tumours. Herein, we report the case of a 59-year-old woman with stage IVB gastric cancer (UICC 8th edition) characterized by FGFR2-TRIM44 fusion and FGFR2 amplification. Her tumour progressed despite multiple lines of standard chemotherapy. Plasma ctDNA analysis revealed these alterations, and pemigatinib was recommended by an expert panel. Treatment led to a remarkable clinical and radiological response, and she remained on pemigatinib with stable symp

Portable detection of Salmonella in food of animal origin via Cas12a-RAA combined with an LFS/PGM dual-signaling readout biosensor.

A highly specific and sensitive rapid two-signal assay was developed for the detection of Salmonella typhimurium in foods of animal origin. The invA gene of Salmonella was used as the biorecognition element and recombinase-assisted amplification (RAA) technology for signal amplification. By utilizing the specific recognition and efficient trans-cleavage activity of CRISPR/Cas12a, point-of-care testing (POCT) for S. typhimurium was achieved via lateral flow strips (LFS) and personal glucometer (PGM) biosensors as dual signal readout systems, with sensitivities of 33 CFU/mL and 20 CFU/mL, respec

Secondary Angiosarcoma With C-MYC Amplification Following Prophylactic Bilateral Mastectomy and Autologous Breast Reconstruction: Report of a Case and Review of the Literature.

In this article, we report a very rare case of secondary angiosarcoma in a young woman with no prior history of breast cancer who had bilateral prophylactic mastectomies with autologous reconstruction due to a strong family history of breast cancer and BRCA1 gene variant of uncertain significance. The surgery was complicated by recurrent fat necrosis requiring several excisions and additional reconstruction followed by the development of localized lymphedema and subsequent angiosarcoma in the reconstructed breast 10 years later. The angiosarcoma was high grade with prominent epithelioid featur

Alu-dependent RNA editing of GLI1 promotes malignant regeneration in multiple myeloma.

Despite novel therapies, relapse of multiple myeloma (MM) is virtually inevitable. Amplification of chromosome 1q, which harbors the inflammation-responsive RNA editase adenosine deaminase acting on RNA (ADAR)1 gene, occurs in 30-50% of MM patients and portends a poor prognosis. Since adenosine-to-inosine RNA editing has recently emerged as a driver of cancer progression, genomic amplification combined with inflammatory cytokine activation of ADAR1 could stimulate MM progression and therapeutic resistance. Here, we report that high ADAR1 RNA expression correlates with reduced patient survival

Совпадение в полном тексте источника
… Gene amplification-associated overexpression of the RNA editing enzyme ADAR1 enhances human lung tumorigenesis. Oncogene 35 , 4407–4013 (2015). 15. Carrasco DR, et al. High-resolution genomic profiles define distinct clinico-pathogenetic subgroups of multiple myeloma patients. …

Microparticle-based RT-qPCR for highly selective rare mutation detection.

The quantitative reverse transcription polymerase chain reaction (RT-qPCR) has become one of the most widely used methods in the detection of disease-specific RNAs. The RT-qPCR involves two separate steps, RT and qPCR. In this study, we suggest a new RT-qPCR protocol with the particles of primer-immobilized networks (PINs), performing capture, RT and amplification of a target RNA in one particle. The production of undesired cDNAs was dramatically suppressed by the specific capture of the target RNA within the particle. Afterward, RT and amplification processes are performed without loss of cDN

Development of internally controlled duplex real-time NASBA diagnostics assays for the detection of microorganisms associated with bacterial meningitis.

Three duplex molecular beacon based real-time Nucleic Acid Sequence Based Amplification (NASBA) assays have been designed and experimentally validated targeting RNA transcripts for the detection and identification of Haemophilus influenzae, Neisseria meningitidis and Streptococcus pneumoniae respectively. Each real-time NASBA diagnostics assay includes an endogenous non-competitive Internal Amplification Control (IAC) to amplify the splice variant 1 mRNA of the Homo sapiens TBP gene from human total RNA. All three duplex real-time NASBA diagnostics assays were determined to be 100% specific fo

LAMP technology: Rapid identification of Brucella and Mycobacterium avium subsp. paratuberculosis.

In this study, we developed new sets of primers to detect Brucella spp. and M. avium subsp. paratuberculosis (MAP) through isothermal amplification. We selected a previously well-characterized target gene, bscp31, specific for Brucella spp. and IS900 for MAP. The limits of detection using the loop-mediated isothermal amplification (LAMP) protocols described herein were similar to those of conventional PCR targeting the same sequences. Hydroxynaphtol blue and SYBR Green(TM) allowed direct naked-eye detection with identical sensitivity as agarose gel electrophoresis. We included the LAMP-based p

Development and application of loop-mediated isothermal amplification for detection of the F167Y mutation of carbendazim-resistant isolates in Fusarium graminearum.

Resistance of Fusarium graminearum to carbendazim is caused by point mutations in the β2-tubulin gene. The point mutation at codon 167 (TTT → TAT, F167Y) occurs in more than 90% of field resistant isolates in China. To establish a suitable method for rapid detection of the F167Y mutation in F. graminearum, an efficient and simple method with high specificity was developed based on loop-mediated isothermal amplification (LAMP). A set of four primers was designed and optimized to specially distinguish the F167Y mutation genotype. The LAMP reaction was optimal at 63 °C for 60 min. When hydroxynap

Evaluation of opa-based real-time PCR for detection of Neisseria gonorrhoeae.

OBJECTIVES: Detection of Neisseria gonorrhoeae by commercial and in-house-based assays has been hampered by false-positive and false-negative results. The current study describes a sensitive and specific real-time 5'-nuclease PCR assay targeting a 90-bp region of the multicopy opa gene. GOAL: To evaluate the sensitivity and specificity of this assay in detection of gonococcus. STUDY: Sensitivity and specificity were determined by testing a panel of 173 microorganisms. In addition, 135 clinical samples previously evaluated by 4 nucleic acid amplification methods were also tested. RESULTS: A

HER-2 and INT-2 amplification estimated by quantitative PCR in paraffin-embedded ovarian cancer tissue samples.

Competitive polymerase chain reaction (PCR) systems were developed for rapid and quantitative estimation of HER-2 (c-erbB-2) and INT-2 oncogene amplification in paraffin-embedded ovarian cancer tissue samples. The beta-globin gene was used as reference and DNA from paraffin-embedded placenta tissue as single copy control. Reliability of the PCR method could be demonstrated by comparing dot blot data with PCR data of identical tumour samples. The PCR method was used to determine HER-2 and INT-2 copy numbers in 196 ovarian cancer samples. HER-2 and INT-2 were found to be amplified in 40 and 19%,…

Diagnosis of typhus infection with Rickettsia tsutsugamushi by polymerase chain reaction.

Two sets of oligonucleotide primers were used to amplify the genomic DNA of Rickettsia tsutsugamushi, the causative agent of scrub typhus (tsutsugamushi disease), by the polymerase chain reaction. Each set of primers amplified 538-bp and 109-bp products, representing part of a gene encoding a possible major 58-kDa immunogenic protein, from whole genomic DNA extracted from R. tsutsugamushi strains Karp, Kato, Gilliam, Kuroki and Kawasaki. No amplification was observed from R. sibirica, R. rickettsii, mouse and human genomic DNA. DNA amplification was observed from crude lysates of peripheral wh

Distribution of p53 codon 72 polymorphism in Indian primary open angle glaucoma patients.

PURPOSE: Glaucoma is a complex neurodegenerative disorder of the eye. Primary Open Angle Glaucoma (POAG) is the most common type, accounting for over half of the total cases. Recently, a significant difference in the distribution of the codon 72 polymorphism of the tumor suppressor gene p53 between control subjects and POAG patients of Chinese origin (p=0.00782) was demonstrated. The proline residue at codon 72 of the p53 gene was significantly over represented in the POAG patients relative to healthy controls. The purpose of this study was to investigate whether the reported association betwe

Targeted Antimicrobial Resistance Gene Screening from Metagenomic DNA of Raw Milk Samples Identifies the Presence of Multiple Genes Including the mcr9.

The current study has investigated the prevalence of antimicrobial resistance (AMR) genes in cow and goat raw milk samples. The misuse of antibiotics in the livestock sector has already been reported to be a major factor contributing to AMR risk. For the study, milk samples were collected from five different farms, and metagenomic DNA was extracted. Then, PCR amplification was carried out using primers specific to 15 different AMR genes. From the results obtained, the prevalence of β-lactam resistance genes, particularly blaTEM (24%), along with other genes like blaZ (12%) and blaSHV (8%), wer

Comparative evaluation of real-time PCR and droplet digital PCR targeting the B646L (p72) and CP204L (p30) genes for early detection of African swine fever virus.

This study compared the analytical performance, diagnostic accuracy, and early detection capability of four nucleic acid amplification assays for African swine fever virus (ASFV), including two real-time PCR (qPCR) assays and two droplet digital PCR (ddPCR) assays targeting the B646L (p72) and CP204L (p30) genes. Analytical performance was evaluated using plasmid standards, whereas diagnostic performance was assessed using 133 samples comprising 73 specimens from experimentally infected pigs and 60 ASFV-negative controls. Both qPCR assays exhibited excellent linearity (R² > 0.99) and acceptabl

GLI1-Altered neoplasms: From molecular alterations to morphologic and phenotypic features with formal recognition in the upcoming WHO Classification of soft tissue tumors.

GLI1-altered mesenchymal tumors represent a recently defined group of neoplasms molecularly characterized by GLI1 gene fusions or amplifications. Although initially thought to share similar molecular alterations, including pericytoma with ACTB::GLI1 fusion, plexiform fibromyxoma, and gastroblastoma, these tumors are now recognized as distinct entities with specific clinicopathological features. Histologically, GLI1-altered mesenchymal tumors typically exhibit a multinodular growth pattern composed of relatively uniform epithelioid-to-ovoid cells arranged in nested formations. These nests are s

TERT Copy Number Alterations, Promoter Mutations and Rearrangements in Adrenocortical Carcinomas.

Molecular characterization of adrenocortical carcinomas (ACC) by The Cancer Genome Atlas (TCGA) has highlighted a high prevalence of TERT al … ailable follow up had adverse outcomes. We found that TERT alterations in ACC predominantly involve gene amplifications, with a smaller subset harboring "hotspot" promoter mutations and rearrangements, and 70% of TERT-altered tumors are associated with metastases. Prospective studies are needed to validate the prognostic impact of these findings.

Development and validation of a whole genome amplification long-range PCR sequencing method for ADPKD genotyping of low-level DNA samples.

Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in two large genes, PKD1 and PKD2, but genetic testing is complicated by the large transcript sizes and the duplication of PKD1 exons 1-33 as six pseudogenes on chromosome 16. Long-range PCR (LR-PCR) represents the gold standard approach for PKD1 genetic analysis. However, a major issue with this approach is that it requires large quantities of genomic DNA (gDNA) material limiting its application primarily to DNA extracted from blood. In this study, we have developed a whole genome amplification (WGA)-based genotyping