Dataset Information

Evaluation of Optimal Reference Genes for qRT-PCR Analysis in Hyphantria cunea (Drury).

ABSTRACT: The relative quantification of gene expression is mainly achieved through reverse transcription-quantitative PCR (qRT-PCR); however, its reliability and precision rely on proper data normalization using one or more optimal reference genes. Hyphantria cunea (Drury) has been an invasive pest of forest trees, ornamental plants, and fruit trees in China for many years. Currently, the molecular physiological role of reference genes in H. cunea is unclear, which hinders functional gene study. Therefore, eight common reference genes, RPS26, RPL13, UBI, AK, RPS15, EIF4A, β-actin, α-tub, were selected to evaluate levels of gene expression stability when subjected to varied experimental conditions, including developmental stage and gender, different tissues, larvae reared on different hosts and different larval density. The geNorm, BestKeeper, ΔCt method, and NormFinder statistical algorithms were used to normalize gene transcription data. Furthermore, the stability/suitability of these candidates was ranked overall by RefFinder. This study provides a comprehensive evaluation of reference genes in H. cunea and could help select reference genes for other Lepidoptera species.

SUBMITTER: Zhao X

PROVIDER: S-EPMC8778541 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Evaluation of Optimal Reference Genes for qRT-PCR Analysis in Hyphantria cunea (Drury).

Zhao Xudong X Geng Yishu Y Hu Tianyi T Zhao Yongang Y Yang Suling S Hao Dejun D

Insects 20220114 1

The relative quantification of gene expression is mainly achieved through reverse transcription-quantitative PCR (qRT-PCR); however, its reliability and precision rely on proper data normalization using one or more optimal reference genes. Hyphantria cunea (Drury) has been an invasive pest of forest trees, ornamental plants, and fruit trees in China for many years. Currently, the molecular physiological role of reference genes in H. cunea is unclear, which hinders functional gene s ...[more]

PMID: 35055939

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Project description:BackgroundBaculoviruses act as effective biological control agents against the invasive pest Hyphantria cunea Drury. In this study, two Chinese Hyphantria cunea nucleopolyhedrovirus (HycuNPV) isolates, HycuNPV-BJ and HycuNPV-HB, were deep sequenced and compared with the Japanese isolate, HycuNPV-N9, to determine whole-genome level diversity and evolutionary history.ResultsThe divergence of the phylogenetic tree and the K2P distances based on 38 core-gene concatenated alignment revealed that two Chinese HycuNPV isolates were a novel species of Alphabaculovirus that infected Hyphantria cunea in China. The gene contents indicated significant differences in the HycuNPV genomes between the Chinese and Japanese isolates. The differences included gene deletions, acquisitions and structural transversions, but the main difference was the high number of single nucleotide polymorphisms (SNPs). In total, 10,393 SNPs, corresponding to approximately 8% of the entire HycuNPV-N9 genome sequence, were detected in the aligned reads. By analyzing non-synonymous variants, we found that hotspot mutation-containing genes had mainly unknown functions and most were early expressing genes. We found that the hycu78 gene which had early and late promoter was under positive selection. Biological activity assays revealed that the infectivity of HycuNPV-HB was greater than that of HycuNPV-BJ, and the killing speed of HycuNPV-HB was faster than that of HycuNPV-BJ. A comparison of molecular genetic characteristics indicated that the virulence differences between the two isolates were affected by SNP and structural variants, especially the homologous repeat regions.ConclusionsThe genomes of the two Chinese HycuNPV isolates were characterized, they belonged to a novel species of Alphabaculovirus that infected Hyphantria cunea in China. We inferred that the loss or gain of genetic material in the HycuNPV-HB and HycuNPV-BJ genomes resulted in new important adaptive capabilities to the H. cunea host. These results extend the current understanding of the genetic diversity of HycuNPV and will be useful for improving the applicability of this virus as a biological control agent.

Dataset Information

Evaluation of Optimal Reference Genes for qRT-PCR Analysis in Hyphantria cunea (Drury).

Publications

Evaluation of Optimal Reference Genes for qRT-PCR Analysis in <i>Hyphantria cunea</i> (Drury).

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