Selection of Reference Genes for MicroRNA Quantitative Expression Analysis in Chinese Perch, Siniperca chuatsi

Zhu, Xin; Li, Yulong; Chen, Dunxue; Wu, Ping; Tan, Yi; Chen, Tao; Zhang, Jianshe; Chu, Wuying

doi:10.3390/ijms16048310

Cited by 20 publications

(22 citation statements)

References 33 publications

(51 reference statements)

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“…The validation of reference genes in our study indicated that three miRNAs, either gmo -17-1—5p alone, or the combination of gmo -miR-25-3p and gmo -miR-210-5p, were particularly suitable as reference genes in Atlantic cod. In agreement with recent finding in similar studies [ 58 ] U6 performed less well compared to the miRNAs as an endogenous control. MiR-25-3p seems to perform well as a miRNA reference gene in several teleosts [ 37 , 40 ].…”

Section: Discussionsupporting

confidence: 93%

Discovery of miRNAs and Their Corresponding miRNA Genes in Atlantic Cod (Gadus morhua): Use of Stable miRNAs as Reference Genes Reveals Subgroups of miRNAs That Are Highly Expressed in Particular Organs

et al. 2016

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BackgroundAtlantic cod (Gadus morhua) is among the economically most important species in the northern Atlantic Ocean and a model species for studying development of the immune system in vertebrates. MicroRNAs (miRNAs) are an abundant class of small RNA molecules that regulate fundamental biological processes at the post-transcriptional level. Detailed knowledge about a species miRNA repertoire is necessary to study how the miRNA transcriptome modulate gene expression. We have therefore discovered and characterized mature miRNAs and their corresponding miRNA genes in Atlantic cod. We have also performed a validation study to identify suitable reference genes for RT-qPCR analysis of miRNA expression in Atlantic cod. Finally, we utilized the newly characterized miRNA repertoire and the dedicated RT-qPCR method to reveal miRNAs that are highly expressed in certain organs.ResultsThe discovery analysis revealed 490 mature miRNAs (401 unique sequences) along with precursor sequences and genomic location of the miRNA genes. Twenty six of these were novel miRNA genes. Validation studies ranked gmo-miR-17-1—5p or the two-gene combination gmo-miR25-3p and gmo-miR210-5p as most suitable qPCR reference genes. Analysis by RT-qPCR revealed 45 miRNAs with significantly higher expression in tissues from one or a few organs. Comparisons to other vertebrates indicate that some of these miRNAs may regulate processes like growth, lipid metabolism, immune response to microbial infections and scar damage repair. Three teleost-specific and three novel Atlantic cod miRNAs were among the differentially expressed miRNAs.ConclusionsThe number of known mature miRNAs was considerably increased by our identification of miRNAs and miRNA genes in Atlantic cod. This will benefit further functional studies of miRNA expression using deep sequencing methods. The validation study showed that stable miRNAs are suitable reference genes for RT-qPCR analysis of miRNA expression. Applying RT-qPCR we have identified several miRNAs likely to have important regulatory functions in particular organs.

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Section: Discussionsupporting

confidence: 93%

Discovery of miRNAs and Their Corresponding miRNA Genes in Atlantic Cod (Gadus morhua): Use of Stable miRNAs as Reference Genes Reveals Subgroups of miRNAs That Are Highly Expressed in Particular Organs

et al. 2016

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“…found that miR-101a was the most stable miRNA when all tissue types were considered separately [ 63 ]. In Siniperca chuatsi , all six miRNAs (miR-101a, miR-146a, miR-22a, miR-23a, miR-26a and let-7a) exhibited better expression consistency than U6 in most of the conditions examined, and the best combination of reference genes was miR-22a and miR-23a [ 64 ]. Herein, we systematically evaluated the reliability of seven miRNA genes across 15 tissues.…”

Section: Discussionmentioning

confidence: 99%

Identification and characterization of skin color microRNAs in Koi carp (Cyprinus carpio L.) by Illumina sequencing

et al. 2018

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BackgroundMicroRNAs (miRNAs) are endogenous, small (21–25 nucleotide), non-coding RNAs that play important roles in numerous biological processes. Koi carp exhibit diverse color patterns, making it an ideal subject for studying the genetics of pigmentation. However, the influence of miRNAs on skin color regulation and variation in Koi carp is poorly understood.ResultsHerein, we performed small RNA (sRNA) analysis of the three main skin colors in Koi carp by Illumina sequencing. The results revealed 330, 397, and 335 conserved miRNAs (belonging to 81 families) and 340, 353, and 351 candidate miRNAs in black, red, and white libraries, respectively. A total of 164 differentially expressed miRNAs (DEMs) and 14 overlapping DEMs were identified, including miR-196a, miR-125b, miR-202, miR-205-5p, miR-200b, and etc. Target prediction and functional analysis of color-related miRNAs such as miR-200b, miR-206, and miR-196a highlighted putative target genes, including Mitf, Mc1r, Foxd3, and Sox10 that are potentially related to pigmentation. Determination of reference miRNAs for relative quantification showed that let-7a was the most abundant single reference gene, and let-7a and miR-26b was the most abundant combination.ConclusionsThe findings provide novel insight into the molecular mechanisms determining skin color differentiation in Koi carp, and serve as a valuable reference for future studies on tissue-specific miRNA abundance in Koi carp.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-5189-5) contains supplementary material, which is available to authorized users.

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“…To validate the Illumina sequencing, 10 conserved miRNAs that have been reported with relative stable expression in other organisms [ 45 – 47 ] and 10 novel identified miRNAs with high abundance in sRNA library of G . molesta ( S1 Table ) were selected and their expression level in the sample for construction of sRNA library were checked using qPCR.…”

Section: Methodsmentioning

confidence: 99%

De novo characterization of microRNAs in oriental fruit moth Grapholita molesta and selection of reference genes for normalization of microRNA expression

Wang

Zhang

et al. 2017

PLoS ONE

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MicroRNAs (miRNAs) are a group of endogenous non-coding small RNAs that have critical regulatory functions in almost all known biological processes at the post-transcriptional level in a variety of organisms. The oriental fruit moth Grapholita molesta is one of the most serious pests in orchards worldwide and threatens the production of Rosacea fruits. In this study, a de novo small RNA library constructed from mixed stages of G. molesta was sequenced through Illumina sequencing platform and a total of 536 mature miRNAs consisting of 291 conserved and 245 novel miRNAs were identified. Most of the conserved and novel miRNAs were detected with moderate abundance. The miRNAs in the same cluster normally showed correlated expressional profiles. A comparative analysis of the 79 conserved miRNA families within 31 arthropod species indicated that these miRNA families were more conserved among insects and within orders of closer phylogenetic relationships. The KEGG pathway analysis and network prediction of target genes indicated that the complex composed of miRNAs, clock genes and developmental regulation genes may play vital roles to regulate the developmental circadian rhythm of G. molesta. Furthermore, based on the sRNA library of G. molesta, suitable reference genes were selected and validated for study of miRNA transcriptional profile in G. molesta under two biotic and six abiotic experimental conditions. This study systematically documented the miRNA profile in G. molesta, which could lay a foundation for further understanding of the regulatory roles of miRNAs in the development and metabolism in this pest and might also suggest clues to the development of genetic-based techniques for agricultural pest control.

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Selection of Reference Genes for MicroRNA Quantitative Expression Analysis in Chinese Perch, Siniperca chuatsi

Cited by 20 publications

References 33 publications

Discovery of miRNAs and Their Corresponding miRNA Genes in Atlantic Cod (Gadus morhua): Use of Stable miRNAs as Reference Genes Reveals Subgroups of miRNAs That Are Highly Expressed in Particular Organs

Discovery of miRNAs and Their Corresponding miRNA Genes in Atlantic Cod (Gadus morhua): Use of Stable miRNAs as Reference Genes Reveals Subgroups of miRNAs That Are Highly Expressed in Particular Organs

Identification and characterization of skin color microRNAs in Koi carp (Cyprinus carpio L.) by Illumina sequencing

De novo characterization of microRNAs in oriental fruit moth Grapholita molesta and selection of reference genes for normalization of microRNA expression

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