Track topics on Twitter Track topics that are important to you
The detection of somatic mutations in the 9 exon of the calreticulin gene (CALR) is regulated by the clinical recommendations as a diagnostic criterion for chronic Ph-negative myeloproliferative neoplasms (MPN). Some methods of nucleic acids testing are used to identify CALR gene mutations with different requirements for special skills of personnel and expensive equipment. The purpose of this work is to compare the results of the detection of CALR gene mutations in venous blood samples by allele-specific RT-PCR with subsequent electrophoresis, fragment analysis and Sanger- or pyro- sequencing. We used 1284 blood samples of patients with suspected MPN and 20 blood donor samples. Mutations in the CALR gene of the I and II type were identified using PCR-RT with the original primers and TaqMan probes. Also, all samples were tested for mutations in the CALR gene by electrophoretic detection of PCR results in an agarose gel. The use of allele-specific RT-PCR followed by electrophoretic detection made it possible to determine clinically significant mutations in the CALR gene in 81 venous blood samples of JAK2- and MPL-negative patients, including 42 cases of type I mutation, 33 cases of type II mutation and 8 rare CALR mutations. Mutations in the 9 exon of the CALR gene were not detected in any of the 20 blood donor samples or in 121 blood samples of patients with polycythemia vera. In randomly selected 20 negative samples, CALR gene mutations were also not detected using Sanger sequencing. All positive samples were confirmed by fragment analysis, as well as with Sanger- sequencing and pyro- sequencing. The described combined approach to detect mutations of the CALR gene in peripheral blood samples can be used in clinical diagnostic laboratories that have a standard set of equipment for electrophoresis of nucleic acids and a PCR-RT. We also propose a confirmatory test based on the pyrosequencing of DNA using the system of genetic analysis "PyroMark Q24".
This article was published in the following journal.
Name: Klinicheskaia laboratornaia diagnostika
To compare the detection results consistency of quantitative polymerase chain reaction (qPCR) and digital droplet polymerase chain reaction (ddPCR), and determine the value of ddPCR for viral detectio...
Incidence of transient bacteremia following dental extractions ranges 30%-70% among adults and 58%-100% in children. This study aims to assess the multiplex polymerase chain reaction (PCR) technique i...
To estimate the clinical value of bacterial detection in peritoneal dialysis-associated peritonitis (PDAP) by multiplex real-time polymerase chain reaction (RT-PCR). This study was undertaken to evalu...
Early detection of tuberculosis is one of the crucial steps for TB control. Although, the sensitivity of conventional methods like Lowenstein Jensen (LJ) culture and direct staining is quite low, mole...
Our study aimed to establish a novel multiplex polymerase chain reaction (PCR) for rapid simultaneous detection of all relevant human neutrophil antigen (HNA)-1, -3, -4 and -5 alleles.
The purpose of this study is to evaluate the added diagnostic value of a quantitative polymerase chain reaction targeting the lytA gene in detecting pneumococci in patients with community-...
The primary purpose of this study is to validate the sensitivity and specificity of the Respirio Flu Test and the eLab Flu Test in detecting Influenza A as compared to the gold standard fo...
The principal objective is to assess the diagnostic accuracy of the PoC assay (Genedrive, Epistem) to detect HCV RNA against the reference standard of commercial real-time polymerase chain...
This study focusses on finding out if osteosarcoma can be detected in blood. The cells will be measured by a new laboratory technique called the polymerase chain reaction. This new techniq...
Group B streptococcus infections may be serious for the neonates. The infection can occur during the birth, by contact with the genital area. That is why the detection of this bacteria is ...
In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships.
Methods for using more than one primer set in a polymerase chain reaction to amplify more than one segment of the target DNA sequence in a single reaction.
Methods used for detecting the amplified DNA products from the polymerase chain reaction as they accumulate instead of at the end of the reaction.
MOLECULAR BIOLOGY techniques used in the diagnosis of disease. Included are such techniques as IN SITU HYBRIDIZATION of chromosomes for CYTOGENETIC ANALYSIS; OLIGONUCLEOTIDE ARRAY SEQUENCE ANALYSIS of gene expression patterns in disease states; identification of pathogenic organisms by analysis of species specific DNA sequences; and detection of mutations with POLYMERASE CHAIN REACTION.
Detection of drugs that have been abused, overused, or misused, including legal and illegal drugs. Urine screening is the usual method of detection.
Polymerase Chain Reaction (PCR)
PCR (Polymerase Chain Reaction) uses the ability of DNA polymerase (enzymes that create DNA molecules by assembling nucleotides, the building blocks of DNA. These enzymes are essential to DNA replication and usually work in pairs to create two ident...
Bioinformatics is the application of computer software and hardware to the management of biological data to create useful information. Computers are used to gather, store, analyze and integrate biological and genetic information which can then be applied...
DNA sequencing is the process of determining the precise order of nucleotides within a DNA molecule. During DNA sequencing, the bases of a small fragment of DNA are sequentially identified from signals emitted as each fragment is re-synthesized from a ...