Development of Sarcophaga dux (Diptera: Sarcophagidae) at constant temperatures and differential gene expression for age estimation of intrapuparial

Background: Sarcophaga dux (Diptera: Sarcophagidae) is a necrophagous flesh fly with potential forensic value in estimating minimum postmortem interval (PMI min ). The basic developmental data and precise intrapuparial age estimates are significant for PMI min estimationof application entomological data in legal medicine. Methods: The development parameters of S. dux at seven constant temperatures from 16°C to 34°C were investigated by rearing using pig lung in the artificial climate box. The appropriate reference genes and intrapuparial differentially expressed genes (DEGs) of S. dux at constant temperatures 34, 25 and 16°C were selected and analyzed using RT-qPCR for more precisely age estimations. Results: The developmental durations of S. dux at 16, 19, 22, 25, 28, 31 and 34°C from larviposition to adult eclosion were 1478.6±18.3, 726.1±15.8, 538.5±0.9, 394.1±9.5, 375.6±10.8, 284.1±7.3, and 252.5±6.1 h, respectively. The thermal summation constant of S. dux was 5341.71±249.29 degree hours, and the developmental threshold temperature was 12.266±0.35°C. The most reliable reference genes under intrapuparial and different temperature in our study were: GST1 and 18S rRNA for 34°C group, GST1 and RPL49 for 25 °C group, and 18S rRNA and 28S rRNA for 16 °C group，the four differential expression genes ( Hsp60 , A-alpha , ARP , RPL8 ) can be used to more precisely intrapuparial age estimation of S. dux . Conclusions: The basic developmental data of S. dux at constant temperatures, such as body length changing of the larva period, accumulated degree hours and duration of development, to establish developmental models that can be used to estimate the PMImin. The selection and evaluation of appropriate reference genes at different experimental conditions are primary with RT-qPCR. The differentially expressed gene can contribute to more accuracy age estimations of S. dux intrapuparial. The result from this study can make contributes to the use of S.dux for estimating PMI min .