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23 July 2020 Evaluation of Reference Genes and Age Estimation of Forensically Useful Aldrichina grahami (Diptera: Calliphoridae) During Intrapuparial Period
Zhuoying Liu, Han Han, Wei Chen, Shiwen Wang, Fanming Meng, Jifeng Cai, Yadong Guo
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Abstract

The minimum postmortem interval (PMImin) could be evaluated from the developmental stage of forensically important insects colonize a corpse, such as blow flies (Diptera: Calliphoridae). Unlike larvae, the developmental stage of which is well established according to their morphology, estimating the age of pupae is proven to be challenging. Recently, several studies reported the regulation of special genes during the development of blow fly pupae. However, gene regulation in Aldrichina grahami during the intrapuparial period remains to be studied. Therefore, we set out to investigate the mRNA levels of heat shock protein 23 (Hsp23), heat shock protein 24 (Hsp24), and 1_16 during the metamorphosis of A. grahami pupae. First, we examined seven candidate reference genes (ribosomal protein 49 (RP49), 18S ribosomal RNA (18S rRNA), 28S ribosomal RNA (28S rRNA), beta-tubulin at 56D-tubulin), Ribosomal protein L23 (RPL23), glutathione S-transferase (GST1), and Actin. Three widely used algorithms (NormFinder, BestKeeper, and geNorm) were applied to evaluate the mRNA levels of reference gene candidates in puparium at three stable temperatures (15, 22, and 27°C). Next, mRNA expression of Hsp23, Hsp24, and 1_16 during A. grahami metamorphosis was examined. We demonstrated that mRNA expression levels of Hsp23, Hsp24, and 1_16 showed time-specific regulation. In summary, our study identified three gene markers for the intrapuparial period of A. grahami and might provide a potential application in PMImin estimation.

© The Author(s) 2020. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
Zhuoying Liu, Han Han, Wei Chen, Shiwen Wang, Fanming Meng, Jifeng Cai, and Yadong Guo "Evaluation of Reference Genes and Age Estimation of Forensically Useful Aldrichina grahami (Diptera: Calliphoridae) During Intrapuparial Period," Journal of Medical Entomology 58(1), 47-55, (23 July 2020). https://doi.org/10.1093/jme/tjaa144
Received: 21 April 2020; Accepted: 16 June 2020; Published: 23 July 2020
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KEYWORDS
Aldrichina grahami
differentially expressed genes
forensic entomology
intrapuparial
postmortem interval
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