Preparing compound heterozygous reference material using gene synthesis technology: a model of thrombophilic mutations

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00179906%3A_____%2F14%3A10283772" target="_blank" >RIV/00179906:_____/14:10283772 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216208:11150/14:10283772

Výsledek na webu

<a href="http://biomed.papers.upol.cz/artkey/bio-201404-0007_preparing_compound_heterozygous_reference_material_using_gene_synthesis_technology_a_model_of_thrombophilic_mu.php#.VK5-h2NWXuc" target="_blank" >http://biomed.papers.upol.cz/artkey/bio-201404-0007_preparing_compound_heterozygous_reference_material_using_gene_synthesis_technology_a_model_of_thrombophilic_mu.php#.VK5-h2NWXuc</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.5507/bp.2014.041" target="_blank" >10.5507/bp.2014.041</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Preparing compound heterozygous reference material using gene synthesis technology: a model of thrombophilic mutations

Popis výsledku v původním jazyce

The aim of our study is to present a novel approach for preparing a compound heterozygous reference material (hetRM) using gene synthesis technology with inverted insertion of wild-type and mutant fragments into a single cloning vector. Factor II (G20210A) and Factor V (G1691A Leiden) gene mutations were used as an experimental model. During the gene synthesis, DNA fragments were aligned in the following order: G1691 FV wild-type forward strain, G20210 FII wild-type forward strain, 1691A FV mutant reverse strain, 20210A FII mutant reverse strain. The complete chain was inserted into a pIDT SMART cloning vector and amplified in an E. coli competent strain. For assessing hetRM characteristics and commutability, we used real-time PCR with subsequent melting curve analysis, real-time PCR with hydrolysis probes, allele-specific amplification, reverse hybridization, and dideoxynucleotide DNA sequencing. All five methods yielded concordant results of DNA analysis of the hetRM. Differences in

Název v anglickém jazyce

Preparing compound heterozygous reference material using gene synthesis technology: a model of thrombophilic mutations

Popis výsledku anglicky

The aim of our study is to present a novel approach for preparing a compound heterozygous reference material (hetRM) using gene synthesis technology with inverted insertion of wild-type and mutant fragments into a single cloning vector. Factor II (G20210A) and Factor V (G1691A Leiden) gene mutations were used as an experimental model. During the gene synthesis, DNA fragments were aligned in the following order: G1691 FV wild-type forward strain, G20210 FII wild-type forward strain, 1691A FV mutant reverse strain, 20210A FII mutant reverse strain. The complete chain was inserted into a pIDT SMART cloning vector and amplified in an E. coli competent strain. For assessing hetRM characteristics and commutability, we used real-time PCR with subsequent melting curve analysis, real-time PCR with hydrolysis probes, allele-specific amplification, reverse hybridization, and dideoxynucleotide DNA sequencing. All five methods yielded concordant results of DNA analysis of the hetRM. Differences in

Druh

J_x - Nezařazeno - Článek v odborném periodiku (Jimp, Jsc a Jost)

CEP obor

EB - Genetika a molekulární biologie

OECD FORD obor

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Projekt

—

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2014

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Biomedical Papers

ISSN

1213-8118

e-ISSN

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Svazek periodika

158

Číslo periodika v rámci svazku

4

Stát vydavatele periodika

CZ - Česká republika

Počet stran výsledku

5

Strana od-do

539-543

Kód UT WoS článku

000347173200007

EID výsledku v databázi Scopus

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