Evaluation of Reference Genes and Age Estimation of Forensically Useful Aldrichina grahami (Diptera: Calliphoridae) During Intrapuparial Period

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 (β-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.

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Evaluation of qPCR reference genes for taimen (Hucho taimen) under heat stress

Scientific Reports ◽

10.1038/s41598-021-03872-x ◽

2022 ◽

Vol 12 (1) ◽

Author(s):

Xiaoxing Yang ◽

Guangxiang Tong ◽

Le Dong ◽

Ting Yan ◽

Huan Xu ◽

...

Keyword(s):

Gene Expression ◽

Heat Stress ◽

Heat Shock ◽

Heat Shock Protein ◽

Ribosomal Protein ◽

Ribosomal Rna ◽

Reference Genes ◽

28S Rrna ◽

Qrt Pcr ◽

Hucho Taimen

AbstractAs a powerful and attractive method for detecting gene expression, qRT-PCR has been broadly used in aquaculture research. Understanding the biology of taimen (Hucho taimen) has drawn increasing interest because of its ecological and economic value. Stable reference genes are required for the reliable quantification of gene expression, but such genes have not yet been optimized for taimen. In this study, the stability levels of 10 commonly used candidate reference genes were evaluated using geNorm, NormFinder, BestKeeper, and RefFinder. The expression levels of the 10 genes were detected using 240 samples from 48 experimental groups consisting of 40 individuals treated under four heat-stress conditions (18, 20, 22, and 24 °C) for 24 h and 26 °C for 4, 24, 48, and 72 h. Six tissues (blood, heart, brain, gill, skin, and liver) were collected from each individual. Ribosomal protein S29 (RPS29) and ribosomal protein L19 (RPL19) were the most stable genes among all of the samples, whereas 28S ribosomal RNA (28S rRNA), attachment region binding protein (ARBP), and 18S ribosomal RNA (18S rRNA) were the least stable. These results were verified by an expression analysis of taimen heat-stress genes (heat shock protein 60, hsp60, and heat shock protein 70, hsp70). In conclusion, RPS29 and RPL19 are the optimal reference genes for qRT-PCR analyses of taimen, irrespective of the tissue and experimental conditions. These results allow the reliable study of gene expression in taimen.

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Brief report: Expression of TNF-α in chronic periapical lesions correlates with expression of bacterial chaperonin-60

Vojnosanitetski pregled ◽

10.2298/vsp201106052s ◽

2021 ◽

pp. 52-52

Author(s):

Jelena Stanisic-Zindovic ◽

Branko Mihailovic ◽

Filip Djordjevic ◽

Marija Milovanovic ◽

Nebojsa Arsenijevic ◽

...

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Mrna Expression ◽

Mrna Levels ◽

Periapical Lesions ◽

Quantitative Expression ◽

Tnf Α ◽

Chaperonin 60 ◽

Anti Inflammatory ◽

Anti Inflammatory Cytokine

Background/Aim: The aim of this study is to determine the quantitative expression of the bacterial heat shock protein, Chaperonin-60 (Cpn60) and pro-inflammatory and anti-inflammatory cytokine in periapical tissue, obtained from individuals with chronic periapical lesions and to determine the correlation between the expression of the bacterial heat shock protein and the expression of these cytokines. Methods. The study was performed on 18 periapical lesions and 6 control samples of healthy periapical tissue, taken at the Clinic of Dental Medicine, Faculty of Medical 4 Sciences University of Pristina, Kosovska Mitrovica. The levels of mRNA expression of pro- and anti- inflammatory cytokines and bacterial heat shock protein were determined by real time quantitative RT-PCR. Results. Analysis revealed significantly higher mRNA levels of TNF-? and Cpn60 in the tissue of periapical lesions compared with normal periapical tissue (P <0.05). Contrary to these results, the mRNA expression of anti-inflammatory IL-10 was significantly higher in the samples of normal periapical tissue compared with the mRNA levels of this cytokine in the tissue of periapical lesions (P <0.001). Expression of Cpn60 is in strong correlation with TNF-? expression in periapical lesions. Conclusion. Cpn60 released from bacteria in periapical tissue could be a strong stimulator of inflammatory response and one of the important players in the pathogenesis of periapical lesions.

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76 EFFECT OF CULTURE METHOD ON THE mRNA EXPRESSION BEFORE AND AFTER CRYOPRESERVATION IN BOVINE BLASTOCYSTS

Reproduction Fertility and Development ◽

10.1071/rdv23n1ab76 ◽

2011 ◽

Vol 23 (1) ◽

pp. 143

Author(s):

A. Kuzmany ◽

V. Havlicek ◽

C. Wrenzycki ◽

S. Wilkening ◽

G. Brem ◽

...

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Mrna Expression ◽

Dna Methyltransferase ◽

Mrna Levels ◽

Solute Carrier Family ◽

Production Methods ◽

Aquaporin 3 ◽

Solute Carrier

Blastocyst mRNA expression and cryopreservability are thought to be suitable indicators of embryo quality and developmental competence and have been shown to be affected by production methods and culture systems. The aim of the present study was to assess cryosurvival and levels of mRNA expression of selected genes [occludin, desmocollin 2, solute carrier family 2 member 3 (formerly glucose transporter 3), BAX, BCL xL, heat shock protein A1A (formerly heat shock protein 70.1), aquaporin 3, and DNA methyltransferase 1a] of bovine blastocysts derived by 4 different, established culture methods [in vitro production (IVP); multiple-ovulation embryo transfer (MOET); transfer into the heifer oviducts of gametes (GIFT); or in vitro derived cleaved stage embryos (Days 2–7)]. Linear models were used for the comparison of the relative abundances of the blastocyst mRNA transcripts. Separate 1-way ANOVA were used. The production methods were used as factors, except for the comparisons between pre- and post-cryopreservation, where 2-way ANOVA were used. The level of significance was set at P ≤ 0.05. A significant difference in re-expansion rates was found only at 24 h post-thawing, with significantly higher rates in blastocysts produced in vitro compared to embryos of the Days 2–7 group. Levels of mRNA expression were assessed using RT-qPCR. Before cryopreservation of embryos, no significant inter-group differences were seen. However, significantly more desmocollin 2 mRNA expression was detected in embryos of the MOET group compared with blastocysts derived by the other production methods. Post-cryopreservation, blastocysts of 3 embryo production groups (IVP, MOET, Days 2–7) were available for analysis. Compared with levels of mRNA expression before cryopreservation, re-expanded blastocysts after cryopreservation showed a significant up-regulation of heat shock protein A1A transcripts in all groups, and of solute carrier family 2 member 3 transcripts only in the IVP-derived group. The BAX, BCL-xL, occludin, and desmocollin 2 were significantly up-regulated in embryos of the MOET and IVP groups after cryopreservation, as compared with their counterparts before cryopreservation. None of the culture groups showed any pre- v. post-cryopreservation differences in the aquaporin 3 and the DNA methyltransferase 1 mRNA levels. Blastocysts derived by transfer of in vitro derived cleaved stage embryos into the oviduct of synchronised heifers (Days 2–7) did not show any pre- v. post-cryopreservation differences in the mRNA levels of any of the assessed genes. These results merit further investigation. After the process of cryopreservation and thawing, re-expanded embryos of the MOET and IVP groups do increase their mRNA levels to prepare for hatching and further development.

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Molecular Cloning and mRNA Expression of Heat Shock Protein Genes and Their Response to Cadmium Stress in the Grasshopper Oxya chinensis

PLoS ONE ◽

10.1371/journal.pone.0131244 ◽

2015 ◽

Vol 10 (7) ◽

pp. e0131244 ◽

Cited By ~ 7

Author(s):

Yuping Zhang ◽

Yaoming Liu ◽

Jianzhen Zhang ◽

Yaping Guo ◽

Enbo Ma

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Mrna Expression ◽

Molecular Cloning ◽

Cadmium Stress ◽

Heat Shock Protein Genes

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Effect of Acute Heat Stress on Heat Shock Protein 70 and Its Corresponding mRNA Expression in the Heart, Liver, and Kidney of Broilers

Asian-Australasian Journal of Animal Sciences ◽

10.5713/ajas.2008.70703 ◽

2008 ◽

Vol 21 (8) ◽

pp. 1116-1126 ◽

Cited By ~ 25

Author(s):

Jimian Yu ◽

Endong Bao

Keyword(s):

Heat Stress ◽

Heat Shock ◽

Heat Shock Protein ◽

Mrna Expression ◽

Heat Shock Protein 70 ◽

Acute Heat Stress ◽

Liver And Kidney

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Identification and Characterization of Three Heat Shock Protein 90 (Hsp90) Homologs in the Brown Planthopper

Genes ◽

10.3390/genes11091074 ◽

2020 ◽

Vol 11 (9) ◽

pp. 1074

Author(s):

Xuan Chen ◽

Ze-Dong Li ◽

Yi-Ting Dai ◽

Ming-Xing Jiang ◽

Chuan-Xi Zhang

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Developmental Stage ◽

Brown Planthopper ◽

Evolutionary Relationship ◽

Heat Shock Protein 90 ◽

Transcript Level ◽

Growth Conditions ◽

Nilaparvata Lugens ◽

Early Developmental Stage

Hsp90 (heat shock protein 90) chaperone machinery is considered to be a key regulator of proteostasis under both physiological and stress growth conditions in eukaryotic cells. The high conservation of both the sequence and function of Hsp90 allows for the utilization of various species to explore new phenotypes and mechanisms. In this study, three Hsp90 homologs were identified in the brown planthopper (BPH), Nilaparvata lugens: cytosolic NlHsp90, endoplasmic reticulum (ER) NlGRP94 and mitochondrial NlTRAP1. Sequence analysis and phylogenetic construction showed that these proteins belonged to distinct classes consistent with the predicted localization and suggested an evolutionary relationship between NlTRAP1 and bacterial HtpG (high-temperature protein G). Temporospatial expression analyses showed that NlHsp90 was inducible under heat stress throughout the developmental stage, while NlGRP94 was only induced at the egg stage. All three genes had a significantly high transcript level in the ovary. The RNA interference-mediated knockdown of NlHsp90 its essential role in nymph development and oogenesis under physiological conditions. NlGRP94 was also required during the early developmental stage and played a crucial role in oogenesis, fecundity and late embryogenesis. Notably, we first found that NlHsp90 and NlGRP94 were likely involved in the cuticle structure of female BPH. Together, our research revealed multifunctional roles of Hsp90s in the BPH.

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