scholarly journals MorphoCatcher: a multiple-alignment based web tool for target selection and designing taxon-specific primers in the loop-mediated isothermal amplification method

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6801
Author(s):  
Fedor V. Shirshikov ◽  
Yuri A. Pekov ◽  
Konstantin A. Miroshnikov
Keyword(s):  
Molecular Diagnostics ◽  
Target Selection ◽  
Primer Design ◽  
Multiple Alignment ◽  
Isothermal Amplification ◽  
Specific Primer ◽  
Bioinformatics Tool ◽  
Loop Mediated Isothermal Amplification ◽  
Amplification Method ◽  
Selection Of

Background Advantages of loop-mediated isothermal amplification in molecular diagnostics allow to consider the method as a promising technology of nucleic acid detection in agriculture and medicine. A bioinformatics tool that provides rapid screening and selection of target nucleotide sequences with subsequent taxon-specific primer design toward polymorphic orthologous genes, not only unique or conserved common regions of genome, would contribute to the development of more specific and sensitive diagnostic assays. However, considering features of the original software for primer selection, also known as the PrimerExplorer (Eiken Chemical Co. LTD, Tokyo, Japan), the taxon-specific primer design using multiple sequence alignments of orthologs or even viral genomes with conservative architecture is still complicated. Findings Here, MorphoCatcher is introduced as a fast and simple web plugin for PrimerExplorer with a clear interface. It enables an execution of multiple-alignment based search of taxon-specific mutations, visual screening and selection of target sequences, and easy-to-start specific primer design using the PrimerExplorer software. The combination of MorphoCatcher and PrimerExplorer allows to perform processing of the multiple alignments of orthologs for informative sliding-window plot analysis, which is used to identify the sequence regions with a high density of taxon-specific mutations and cover them by the primer ends for better specificity of amplification. Conclusions We hope that this new bioinformatics tool developed for target selection and taxon-specific primer design, called the MorphoCatcher, will gain more popularity of the loop-mediated isothermal amplification method for molecular diagnostics community. MorphoCatcher is a simple web plugin tool for the PrimerExplorer software which is freely available only for non-commercial and academic users at http://morphocatcher.ru.

In-Silico Automated Allele-Specific Primer Design for Loop-Mediated Isothermal Amplification

2020 ◽  
Author(s):  
George Alexandrou ◽  
Jesus Rodriguez-Manzano ◽  
Kenny Malpartida-Cardenas ◽  
Pantelis Georgiou ◽  
Chris Toumazou ◽  
...  
Keyword(s):  
In Silico ◽  
Primer Design ◽  
Isothermal Amplification ◽  
Specific Primer ◽  
Loop Mediated Isothermal Amplification ◽  
Allele Specific

scholarly journals Evaluation of a Loop-Mediated Isothermal Amplification Assay to Detect Carbapenemases Directly From Bronchoalveolar Lavage Fluid Spiked With Acinetobacter spp.

2021 ◽  
Vol 11 ◽  
Author(s):  
Javier Moreno-Morales ◽  
Andrea Vergara ◽  
Tomislav Kostyanev ◽  
Jesús Rodriguez-Baño ◽  
Herman Goossens ◽  
...  
Keyword(s):  
Bronchoalveolar Lavage ◽  
Molecular Diagnostics ◽  
Bronchoalveolar Lavage Fluid ◽  
Lavage Fluid ◽  
Isothermal Amplification ◽  
Loop Mediated Isothermal Amplification ◽  
Carbapenem Resistant ◽  
Amplification Assay ◽  
Acinetobacter Spp ◽  
Selection Of

Carbapenem-resistant Acinetobacter spp. mainly Acinetobacter baumannii are frequently causing nosocomial infections with high mortality. In this study, the efficacy of the Eazyplex® SuperBug Complete A system, based on loop-mediated isothermal amplification (LAMP), to detect the presence of carbapenemases in Acinetobacter spp. directly from bronchoalveolar lavage (BAL) samples was assessed. A total of 22 Acinetobacter spp. strains producing OXA-23, OXA-40, OXA-58, NDM, and IMP were selected. Eazyplex SuperBug Complete A kit, used with the Genie II device, is a molecular diagnostics kit that detects a selection of genes that express carbapenemases (blaKPC, blaNDM, blaVIM, blaOXA–48, blaOXA–23, blaOXA–40, and blaOXA–58). Negative BAL samples were identified, McFarland solutions were prepared from each of the 22 Acinetobacter strains and serial dilutions in saline solution were made to finally spike BAL samples to a concentration of 102 and 103 CFU/ml. Fifteen concentrations out of the 44 tested out did not provide detection of the carbapenemase-producing gene, all but one being at the lowest concentration tested at 102 CFU/ml; therefore, the limit of sensitivity is 103 CFU/ml. This assay represents the kind of advantages that investing in molecular diagnostics brings to the clinical practice, allowing the identification of carbapenemases in less than 30 min with a sensitivity of 103 CFU/ml.

scholarly journals P02.06 A Novel Loop-Mediated Isothermal Amplification Method for Robust Detection of EGFR Mutations

2021 ◽  
Vol 16 (3) ◽  
pp. S248-S249
Author(s):  
Y. Saito ◽  
A. Matsui ◽  
S. Michiyuki ◽  
Y. Yamauchi ◽  
N. Takahashi ◽  
...  
Keyword(s):  
Isothermal Amplification ◽  
Egfr Mutations ◽  
Robust Detection ◽  
Loop Mediated Isothermal Amplification ◽  
Amplification Method

scholarly journals Sensitive and Rapid Detection of Edwardsiellosis in Fish by a Loop-Mediated Isothermal Amplification Method

2004 ◽  
Vol 70 (1) ◽  
pp. 621-624 ◽  
Author(s):  
Ram Savan ◽  
Arisa Igarashi ◽  
Satoru Matsuoka ◽  
Masahiro Sakai
Keyword(s):  
Rapid Detection ◽  
Japanese Flounder ◽  
Isothermal Amplification ◽  
Edwardsiella Tarda ◽  
Fish Disease ◽  
Fish Pathogen ◽  
Loop Mediated Isothermal Amplification ◽  
Diagnostic Protocol ◽  
Amplification Method ◽  
Hemolysin Gene

ABSTRACT Here we report a rapid and sensitive method (using loop-mediated isothermal amplification [LAMP]) for the diagnosis of edwardsiellosis, a fish disease caused by Edwardsiella tarda, in Japanese flounder. A set of four primers was designed, and conditions for the detection were optimized for the detection of E. tarda in 45 min at 65°C. No amplification of the target hemolysin gene was detected in other related bacteria. When the LAMP primers were used, detection of edwardsiellosis in infected Japanese flounder kidney, and spleen and seawater cultures was possible. We have developed a rapid and sensitive diagnostic protocol for edwardsiellosis detection in fish. This is the first report of the application of LAMP for the diagnosis of a fish pathogen.

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