Expression of mapk1 and egr1 genes in Onchidium reevesii under tidal stimulation

Previous studies have speculated that tidal rhythm of Onchidium reevesii was linked to its memory ability, which has not been well explained. Mapk1 and egr1 genes are closely related to memory formation in the MAPK signaling pathway that have been extensively studied in vertebrates. These two genes are involved in Long-term potentiation (LTP), which is generally regarded as one of the main molecular mechanisms underlying learning and memory. To investigate the relationship between tidal rhythm of O. reevesii and its memory ability, we studied the molecular mechanism of mapk1 and egr1 genes cooperating with tidal rhythm. The full-length cDNA sequence of mapk1 gene was cloned by RACE method and analyzed by bioinformatics, then qRT-PCR was used to analyze its expression levels in each tissue. A homology analysis and phylogenetic tree reconstruction revealed that O. reevesii is most closely related to the Biomphalaria glabrata. The qRT-PCR showed that mapk1 was expressed in all the tissues, but the highest expression was in the ganglion. We speculated that mapk1 is related to memory formation of O. reevesii. Then we used qRT-PCR to detect the expression of mapk1 and egr1 in ganglion of O. reevesi under tidal stimulation. The expression of mapk1 and egr1 at the rising tide points were significantly higher (P < 0.05) than that at the previous one lowest tide points on May 15-21, except the rising tide point on May 20 and May 21 of mapk1. The expression of mapk1 and egr1 genes in the ganglion sampled for 7-day were basically the same as that of the tide change of the same day. The expression of the two genes were upregulated at raising tide and downregulated at the lowest tide, it is presumed that the O. reevesii was affected by the tide for a long time and formed the tide memory.

Determining the biological zero for gonadal development razor clams Sinonovacula constricta (lamarck 1818) in Zhejiang,China

The biological zero of gonadal development and the effective accumulated temperature of one and two-year-old razor clams Sinonovacula constricta(Lamarck 1818)were studied in Sanmen County,Zhejiang Province,China between April 2016 to October 2017,using a new variety named“Shen-Zhe No.1”.The biological zero was determined by using the rule of effective accumulated temperature and the linear regression method,which was confirmed using a histological analysis of the full gonadal development cycle.The results showed that the biological zero of gonadal reproduction of 1-year-old and 2-year-old S.constricta was the same:20.8℃.When the temperature reached 20–21℃,analysis of the histological sections indicated that more than half of the S.constricta individuals initiated gonadal development.The average effective accumulated temperature over two years was 806 d℃.This study contributes to our understanding of the regulation and control of gonadal maturation,suggesting that changes in the spawning and breeding season during one-year of farming using the new variety of razor clam could provide an economic benefit to industrialized large-scale spat production.The results demonstrated that the razor clam S.constricta is a typical clam of the accumulative temperature mature oviposition type.In addition,although S.constricta does not display hypothermy,it requires a lower temperature for gonadal maturity and spawning.

Analyses of the differential expression of cloned Caspase-3 and Caspae-9 genes,of Onchidium reevesii,exposed to extreme hot and cold conditions

During low tide intertidal organisms face severe temperature fluctuations.This situation may be more pronounced in the context of climate change.To investigate the molecular response mechanism of Onchidium reevesii when stressed by hot and cold temperatures,we cloned its Caspase-3 and Caspase-9 genes and detected their expression characteristics using RT-qPCR.The results showed that the full-length of the cDNA of Caspase-3 consist of 1831 bp,and included an open reading frame(ORF)of 894 bp,a 50UTR of 155 bp and a 30 UTR of 300 bp,and encoding for 297 amino acids.The full-length of the cDNA of Caspase-9 consisted of 2425 bp,and included an ORF of 1373 bp,a 50 UTR of 290 bp,and a 30 UTR of 759 bp,and encoding for 457 amino acids.During the evolution of species the two genes have been greatly conserved.A homology analysis and phylogenetic tree reconstruction revealed that O.reevesii is most closely related to Aplysia californica.The RT-qPCR showed that both genes were expressed in all of the tissues and that expression was highest in the hepatopancreas.It is speculated that these genes play an essential role in the immunity of O.reevesii to certain diseases.Conditions of temperature stress(40�C and 10�C)showed that both heat and cold stresses could induce significant changes in both genes.Compared to the control group,under cold stress conditions the levels of expression of Caspase-3 and Caspase-9 were significantly upregulated(P<0.05)in each experimental group.When exposed to a constant temperature of 10�C,their expression peaked after 1 h(4.32-fold,P<0.05;and 5.58-fold,P<0.05,respectively)and was at its lowest after 12 h(1.93-fold,P<0.05;and 2.35-fold,P<0.05,respectively).Their expression was upregulated after 24 h(2.68-fold,P<0.05;and 4.21-fold,P<0.05,respectively).The expression of both genes showed similar trends.Under heat stress,the expressions of the two genes showed a wave pattern.When exposed to a constant temperature of 40�C,their expression was up-regulated after 1,6,and 24 h,and down-regulated after 3 and 12 h.Except at 24 h,the other experimental groups did not differ significantly(P>0.05)from the control group.When subjected to various stress conditions the two genes displayed different expression patterns,indicating that the species has different strategies for coping with temperature changes.It further indicates that the effect of temperature on the species may play a role through the Caspase-dependent pathway.These may help to better understand the response,at the molecular level,of O.reevesii to conditions of hot and cold stress.