Abstract
Despite scientific progress, the gene sequences for many species not commonly used in research have not yet been analyzed. This makes it difficult to carry out molecular studies on such animals, as the sequence of genes is the basic information used in many techniques. In this study, we attempt to design primers for a real-time PCR analysis, basing on a comparative analysis of selected gene sequences of species related to Reeves’s muntjac (Muntiacus reevesi) and by identifying highly conservative regions. Results of PCR products sequencing and their alignment with the GenBank collection show that all selected primers gave products highly similar (> 90%) to the intended target (among compared species), which led us to the conclusion that our primers may be used for further analyses of gene expression.
Highlights
Real-time PCR is a basic method for a gene expression analysis and is widely used by scientists all over the world [1]
The specific aim of this study is to design and validate primers that will be used for determining the mRNA expression of eleven genes in the tissue samples collected from the gastrointestinal tract (GIT) of Reeves’s muntjac (Muntiacus reevesi)
The query coverage for analyzed PCR products with target gene sequences of compared species was high for the majority of the primers (≥ 90%)
Summary
Real-time PCR is a basic method for a gene expression analysis and is widely used by scientists all over the world [1]. Primers are designed on the basis of sequences available in GenBank—a collection of publicly available DNA sequences. The genomes of relatively few animal species have been fully sequenced and many available sequences, especially for species not commonly used in research, are partial or putative. The aim of this study is to develop a method of designing primers for a selected gene expression analysis, using real-time PCR, when there is no sequence data available in GenBank for the particular animal. The specific aim of this study is to design and validate primers that will be used for determining the mRNA expression of eleven genes in the tissue samples collected from the gastrointestinal tract (GIT) of Reeves’s muntjac (Muntiacus reevesi)
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