Identification of normalization factors for quantitative real-time RT-PCR analysis of gene expression in Pacific abalone Haliotis discus hannai

Reng Qiu; Boguang Sun; Shasha Fang; Li Sun; Xiao Liu

doi:10.1007/s00343-013-2221-0

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Identification of normalization factors for quantitative real-time RT-PCR analysis of gene expression in Pacific abalone Haliotis discus hannai

Updated：2023-05-16

- Identification of normalization factors for quantitative real-time RT-PCR analysis of gene expression in Pacific abalone Haliotis discus hannai
- Journal of Oceanology and Limnology Vol. 31, Issue 2, Pages: 421-430(2013)
- Affiliations：
  
  1. Institute of Oceanology Chinese Academy of Sciences
  2. Graduate University of Chinese Academy of Sciences
- Author bio：
- Funds：
- DOI：10.1007/s00343-013-2221-0
  CLC：
- Online First：07 March 2013，
  
  Published：2013
- Accepted：
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Qiu, R., Sun, B., Fang, S. et al. Identification of normalization factors for quantitative real-time RT-PCR analysis of gene expression in Pacific abalone Haliotis discus hannai, ., Chin. J. Ocean. Limnol. 31, 421–430 (2013). https://doi.org/10.1007/s00343-013-2221-0
DOI：

Qiu, R., Sun, B., Fang, S. et al. Identification of normalization factors for quantitative real-time RT-PCR analysis of gene expression in Pacific abalone Haliotis discus hannai, ., Chin. J. Ocean. Limnol. 31, 421–430 (2013). https://doi.org/10.1007/s00343-013-2221-0 DOI：

摘要

Abstract

Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) is widely used in studies of gene expression. In most of these studies

housekeeping genes are used as internal references without validation. To identify appropriate reference genes for qRT-PCR in Pacific abalone Haliotis discus hannai

we examined the transcription stability of six housekeeping genes in abalone tissues in the presence and absence of bacterial infection. For this purpose

abalone were infected with the bacterial pathogen Vibrio anguillarum for 12 h and 48 h. The mRNA levels of the housekeeping genes in five tissues (digestive glands

foot muscle

gill

hemocyte

and mantle) were determined by qRT-PCR. The PCR data was subsequently analyzed with the geNorm and NormFinder algorithms. The results show that in the absence of bacterial infection

elongation factor-1-alpha and beta-actin were the most stably expressed genes in all tissues

and thus are suitable as cross-tissue type normalization factors. However

we did not identify any universal reference genes post infection because the most stable genes varied between tissue types. Furthermore

for most tissues

the optimal reference genes identified by both algorithms at 12 h and 48 h post-infection differed. These results indicate that bacterial infection induced significant changes in the expression of abalone housekeeping genes in a manner that is dependent on tissue type and duration of infection. As a result

different normalization factors must be used for different tissues at different infection points.

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references

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