quantitative rt-pcr based platform for rapid quantification of the transcripts of highly homologous multigene families and their members during grain development定量rt - pcr为基础平台的快速量化记录高度同源基因的家庭及其成员在粮食发展.pdfVIP
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quantitative rt-pcr based platform for rapid quantification of the transcripts of highly homologous multigene families and their members during grain development定量rt - pcr为基础平台的快速量化记录高度同源基因的家庭及其成员在粮食发展
Kaczmarczyk et al. BMC Plant Biology 2012, 12:184
/1471-2229/12/184
METHODOLOGY ARTICLE Open Access
Quantitative RT-PCR based platform for rapid
quantification of the transcripts of highly
homologous multigene families and their
members during grain development
1 2 3 1*
Agnieszka Kaczmarczyk , Steve Bowra , Zoltan Elek and Eva Vincze
Abstract
Background: Cereal storage proteins represent one of the most important sources of protein for food and feed
and they are coded by multigene families. The expression of the storage protein genes exhibits a temporal
fluctuation but also a response to environmental stimuli. Analysis of temporal gene expression combined with
genetic variation in large multigene families with high homology among the alleles is very challenging.
Results: We designed a rapid qRT-PCR system with the aim of characterising the variation in the expression of
hordein genes families. All the known D-, C-, B-, and γ-hordein sequences coding full length open reading frames
were collected from commonly available databases. Phylogenetic analysis was performed and the members of the
different hordein families were classified into subfamilies. Primer sets were designed to discriminate the gene
expression level of whole families, subfamilies or individual members. The specificity of the primer sets was
validated before successfully applying them to a cDNA population derived from developing grains of field grown
Hordeum vulgare cv. Barke. The results quantify the number of moles of transcript contributed to a particular gene
family and its subgroups. More over the results indicate the genotypic specific gene expression.
Conclusions: Quantitative RT-PCR with SYBR Green labelling can be a useful technique to follow gene expressio
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