| Hucho taimen grow fast, eat delicious, is one of the rare species of cold waterfishes, it has high economic values.The farmed Hucho taimen culture constantlyexpansion of the scale, the demand for hucho taimen seedling increased, seedlingmortality has become one of the factors that restrict the development of artificialbreeding of hucho taimen. Therefore, the research on immunity and reproductivedevelopment is urgent need at the early stages of hucho taimen. In this study, wedetected the enzyme activity, testosterone and estradiol of hucho taimen in earlydevelopmental stages, and vasa gene cloning and expression analysis, the results are asfollows:1At early development stage, from embryonic to larvae stage, we detected alkalinephosphatase (AKP), acid phosphatase (ACP), superoxide dismutase (SOD), catalase(CAT) activity and glutathione (GSH) and malondialdehyde (MDA) content tounderstand the changes of immune enzymes activity and antioxidant contents of fishbody, we found that at embryos ACP activity was significantly lower than that inadulthood[1],in the embryonic period ACP did not play an important role. In thehatching, however, ACP activity was significantly increased, suggesting that ACP playsan important role in the membrane phase of hucho taimen. In the developing phase ofthe larvae, the activity of ACP has been maintained at a high level to protect fishes. Butthe activity of AKP was significantly lower than that of ACP. In the embryonic period,the activity of AKP is low and declining, only from the hatching to30days later there isa transient increase then down again, show that AKP is not important in the embryonicperiod, temporary improvement only in the hatching phase, which may result the weakof fish digestive absorption, the difficult of absorption of amino acids and othermacromolecular substance, speculate that this may be the cause of hucho taimenseedling growth speed low[2]. In the embryonic and larval and juvenile period, SODactivity had no significant change, has been maintained at between1.46-1.77U, showthat at early developmental stage, SOD did not play a major role in antioxidant.Similarly, CAT activity is very low, almost negligible, show that at early developmentalstage, CAT hardly exert antioxidant effects. At hatching, GSH contents increased,reaching a maximum value of3.7μ mol/gprot, shows that at hatching, GSH play animportant antioxidant role, promote fish growth. In the larval development period,although the contents of GSH is lower than the contents of hatching phase, but remainedat a high level, indicating that GSH in hucho taimen from hatching to the larvae stage play an important antioxidant role, to protect fishes. In the embryonic period, the valueof MDA is at0.1nmol/mgprot then floating up and down, the contents is very low,showed that at this period, peroxidation of hucho taimen is very low. But from thehatching phase to the the juvenile stage, MDA contents constantly increased, showedthat after hatching period, lipid peroxidation of hucho taimen increased significantly,during the larval development, the degree of the attack of free radicals to body cellsincreased significantly.2At the early stage of hucho taimen, the contents of estradiol (E2) is high in vivo,but has a great fluctuation. From the early embryos to the stage before hatching, thecontents of E2constantly increased to the peak, but at hatching phase, the contents ofE2decreased to a lower level. Later, in the larval stage, the E2contents began to riseagain, until the juvenile stage, then decreased, and it was significantly lower than in theembryo and larval stage. And testosterone (T) contents, in the embryonic period,continued to rise to the highest point, until hatching, significantly declined to the lowestpoint, at the larvae stage, the contents of T increased slightly and remained stable trend.It indicated that E2and T contents in embryonic period were higher than in the larvaland juvenile period. At hatching stage, E2and T contents were significantly decreased,while T contents falled to low levels, and then went up slightly and remained stable, E2was significantly increased to the peak at the larval development period,and then falledagain at young fish stage.3In this research, we clone vasa gene, the DNA sequencing results showed that,the full-length of vasa gene was2711bp in hucho taimen (GenBank ID:KF554113),gene open reading frame (ORF) is1974bp long, encoding658amino acids. The aminoacid sequence of DEAD protein contains all of the nine conserved domains:xYxxPTPVQ, AQTGSGKTA, PTRELINQ, TPGR, DEAD, SAT, RGLD, HRIGRTGRand GG repeat sequence[3]. Compared the sequences with other species’ known fromGenBank nucleic acid database, vasa gene in hucho taimen was found similar to that inSalmo salar.Compared with Salmo salar, Oncorhynchus mykiss and Salvelinus pluviusHilgendorf, the similarity are respectively93%,92%and91%. It shows that, vasa genesequences are quite conservative in salmonidae.Through RT-PCR analysis and semi-quantification real-time PCR analysis, in oneage hucho taimen, vasa gene expressed only in testis and ovaries, which is consistentwith previous reports, the results showed that the vasa gene is specifically expressed in germ cell line, it can be used as a molecular marker tracking primordial germ cells(PGCs) formation, migration, differentiation, help to understand the mechanism ofgonadal sex differentiation. At the same time, the expression of vasa gene in the ovariesis significantly higher than that of testis. This is consistent with the previously reportedin other invertebrates and vertebrates in the study, indicated that vasa mRNA is thematernally derived. In different periods of hucho taimen embryos, sustained expressionof vasa gene in early embryos, such as the8cell stage was at the highest expressionlevel, then declined, from differentiation stage to hatching, the expression level of vasagene decreased to a low level, also confirmed that the gene is maternally derived. |
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