High‐resolution melting analysis for detection of fusion allele of <i>FUT2</i>

Soejima, Mikiko; Koda, Yoshiro

doi:10.1002/elps.202000241

Cited by 7 publications

(3 citation statements)

References 18 publications

(32 reference statements)

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“…High resolution melting (HRM) technology plays a vital role in genotyping (Li et al, 2019;Soejima and Koda, 2021). Through HRM and sequencing analyses, we determined that all tfoX genes can be divided into two types.…”

Section: Discussionmentioning

confidence: 99%

The molecular diversity of transcriptional factor TfoX is a determinant in natural transformation in Glaesserella parasuis

et al. 2022

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Natural transformation is a mechanism by which a particular bacterial species takes up foreign DNA and integrates it into its genome. The swine pathogen Glaesserella parasuis (G. parasuis) is a naturally transformable bacterium. The regulation of competence, however, is not fully understood. In this study, the natural transformability of 99 strains was investigated. Only 44% of the strains were transformable under laboratory conditions. Through a high-resolution melting curve and phylogenetic analysis, we found that genetic differences in the core regulator of natural transformation, the tfoX gene, leads to two distinct natural transformation phenotypes. In the absence of the tfoX gene, the highly transformable strain SC1401 lost its natural transformability. In addition, when the SC1401 tfoX gene was replaced by the tfoX of SH0165, which has no natural transformability, competence was also lost. These results suggest that TfoX is a core regulator of natural transformation in G. parasuis, and that differences in tfoX can be used as a molecular indicator of natural transformability. Transcriptomic and proteomic analyses of the SC1401 wildtype strain, and a tfoX gene deletion strain showed that differential gene expression and protein synthesis is mainly centered on pathways related to glucose metabolism. The results suggest that tfoX may mediate natural transformation by regulating the metabolism of carbon sources. Our study provides evidence that tfoX plays an important role in the natural transformation of G. parasuis.

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Section: Discussionmentioning

confidence: 99%

The molecular diversity of transcriptional factor TfoX is a determinant in natural transformation in Glaesserella parasuis

et al. 2022

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“…For this purpose, we recently developed several HRM-based real-time PCR methods for detection of se fus , se 428 , and se 385 alleles 15,20,21 . Predominant se alleles in several South Asian populations are se 428 , se 302 , and se del11 .…”

Section: Discussionmentioning

confidence: 99%

Rapid detection of sedel and 302C>T (rs200157007) responsible for nonsecretor phenotype by real-time PCR-based methods

Soejima

Koda

2021

Preprint

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The sedel allele is one of the nonsecretor alleles (se) of FUT2 generated by an Alu-mediated recombination event and was first found in an Indian Bombay phenotype. Like sedel, se302 having a missense single nucleotide polymorphism (SNP), 302C > T, is characteristic of South Asians with a frequency of 10–30%. We developed a real-time PCR melting curve analysis for detection of sedel using a 127-bp amplicon encompassing the breakpoint junction. In addition, by performing duplex PCR by amplifying a 65-bp amplicon of the FUT2 coding region at the same time, we could determine the zygosity of sedel in a single tube. We also developed an Eprobe-mediated PCR assay (Eprobe-PCR) for detection of 302C > T of FUT2. These methods were validated by analyzing 58 Tamils and 54 Sinhalese in Sri Lanka. Both the duplex PCR melting curve analysis for determination of sedel zygosity and the Eprobe-PCR assay for detection of 302C > T exactly determined three genotypes. In addition, the results of the present methods were in complete agreement with those obtained by previously established methods. The two present methods were reliable and seem to be advantageous for large-scale association studies of FUT2 polymorphisms in South Asian populations.

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“…However, this allele was judged to be the A allele of 385A>T by present probe genotyping and the C allele of 357C>T and A of 385A>T by probe and amplicon genotyping. Previously, we developed the hydrolysis probe (TaqMan) assay and HRM assay for detection of se fus [17,18]. In addition, se alleles having 571C>T (rs1800028), 628C>T (rs1800029), and 849G>A (rs1800030) were codistributed with 385A>T in several East and Southeast Asian populations, such as Filipino, Taiwan aborigines, and Polynesians, with relatively high frequencies (1-20%) [19][20][21].…”

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Detection of the weak‐secretor rs1047781 (385A>T) single nucleotide polymorphism using an unlabeled probe high‐resolution melting‐based method

Soejima

Koda

2021

Electrophoresis

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FUT2 encodes galactoside 2-α-l-fucosyltransferase 2 which determines the secretor status of ABO(H) blood group antigens. Secretors have at least one functional FUT2 allele (Se), while nonsecretors or weak secretors are homozygous for nonfunctional (non-or weak secretor) FUT2 alleles (se or Se w ). The Se w having the 385A>T missense SNP (rs1047781) is the prevalent nonfunctional allele in East and Southeast Asians. In this study, we developed an unlabeled-probe high-resolution melting (HRM) analysis for genotyping of 385A>T and validated the method by analyzing 72 Japanese whose 385A>T genotypes were confirmed by DNA sequencing. The unlabeled-probe HRM analysis clearly discriminated three genotypes of 385A>T. In addition, the results obtained for the 72 Japanese by this method were fully concordant with previous ones. Estimation of secretor status using this cost-effective method may be useful in East and Southeast Asian populations.

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High‐resolution melting analysis for detection of fusion allele of FUT2

Cited by 7 publications

References 18 publications

The molecular diversity of transcriptional factor TfoX is a determinant in natural transformation in Glaesserella parasuis

The molecular diversity of transcriptional factor TfoX is a determinant in natural transformation in Glaesserella parasuis

Rapid detection of sedel and 302C>T (rs200157007) responsible for nonsecretor phenotype by real-time PCR-based methods

Detection of the weak‐secretor rs1047781 (385A>T) single nucleotide polymorphism using an unlabeled probe high‐resolution melting‐based method

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High‐resolution melting analysis for detection of fusion allele of FUT2

Cited by 7 publications

References 18 publications

The molecular diversity of transcriptional factor TfoX is a determinant in natural transformation in Glaesserella parasuis

The molecular diversity of transcriptional factor TfoX is a determinant in natural transformation in Glaesserella parasuis

Rapid detection of sedel and 302C&gt;T (rs200157007) responsible for nonsecretor phenotype by real-time PCR-based methods

Detection of the weak‐secretor rs1047781 (385A>T) single nucleotide polymorphism using an unlabeled probe high‐resolution melting‐based method

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Rapid detection of sedel and 302C>T (rs200157007) responsible for nonsecretor phenotype by real-time PCR-based methods