环境和营养因子对四倍体栉孔扇贝胚胎及早期幼虫的影响
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摘要
本文以栉孔扇贝(Chlamys ferreri)为研究对象,利用细胞松弛素B(CB)抑制栉孔扇贝受精卵第一极体(PB1)释放,获得四倍体材料。研究在不同环境和营养条件下,药物诱导后的栉孔扇贝胚胎及早期幼虫的发育情况,以及早期幼虫倍化率的变化情况。
本实验总共设2个环境因子和5个营养因子:温度、盐度、谷氨酸、牛磺酸、谷氨酰胺、葡萄糖、维生素C。通过统计各实验组的囊胚、原肠胚、担轮幼虫、畸形幼虫及滞育幼虫的比例,以及检测担轮幼虫和D形幼虫两个时期的幼虫倍性组成,来揭示四倍体栉孔扇贝胚胎及早期幼虫的一些生理特性。结果显示,从囊胚到担轮幼虫,处理组均以20℃组发育最佳,而进入孵化期后,22℃组的孵化率显著高于其他组,这说明了处理组存在着温度补偿,适当地提高温度可以促进四倍体胚胎的孵化。同时发现诱导后的四倍体胚胎对温度和盐度的变化比较敏感,最适盐度和温度范围均缩小了,分别为18-22℃和盐度30-34。而从添加营养因子的实验结果来看:胚胎发育过程中对谷氨酸需求量越来越大,谷氨酸的最佳添加量由囊胚时期的0.5μmol/L上升至孵化时期的2.5μmol/L;在囊胚至担轮幼虫的发育过程中40μmol/L浓度的谷氨酰胺可以显著促进胚胎的发育;10 mmol/L的牛磺酸则可以显著提高囊胚率,但是随着胚胎的进一步发育,其对牛磺酸的需求越来越小,至担轮幼虫时期只有5mmol/L浓度的牛磺酸起作用积极的作用;葡萄糖对囊胚的发育没有积极的帮助,而进入原肠胚时期时,胚胎对葡萄糖浓度的需量逐渐增加,到了担轮幼虫时期,2mmol/L浓度的葡萄糖可以大大提高担轮幼虫的比例,显著降低胚胎的畸形率和滞育率;0.009g/L浓度的维生素C显著促进囊胚的发育,而高浓度的维生素C则抑制胚胎的发育,进入原肠胚时,胚胎对维生素C的需求量减少了,当胚胎发育至担轮幼虫时期时,维生素C添加组除了0.003g/L组外,其他处理组均比对照组低。
本研究初步探明四倍体栉孔扇贝胚胎及早期幼虫的发育情况,以及其对环境和营养的需求情况,首次报道了栉孔扇贝四倍体诱导后的胚胎幼虫存在温度补偿和营养补偿。这将为攻克四倍体倍体育种中出现的高死亡率的问题提供解决方案,对贝类四倍体技术的完善具有重要的参考价值。
In this study, we tested tetraploid induction in Chlamys ferreri by inhititing the release of polar body I(PBl)in newly feitilized eggs. To produce tetraploid,fertilized eggs of Chlamys ferreri were treated with Cytochalasin B (CB) to inhibit the release of the PB I .The development and ploidy composition of induced embryo and early larvae were studied in different circumstance and nutrition's factor.
This study was divided into 7 parts, there were: Temperature,Salinity, Glutamic acid,Taurine,GIutamine,Glucose,Vitamine C. By count the change of the blastula rate, gastrulae rate, hatching rate,trochophore-stage larvae rate, abnormal larvae rate, developmentarrested larvae rate, and the ploidy composition of trochophore-stage and D-stage larvae to open out some traits of physiology on embryo and early larvae of tetraploid Chlamys ferreri .the result show that :from the blastula to trochophore-stage larvae ,20°C was the best temperature to develop the embryo in all treated group.But when the trochophore stage larcae become to hatch,the best temperature was ascended to 22°C ,this explain that there had temperature compensation during the development of ploidy embryo and early larvae.The induced embryo and early larvae were susceptivity to the change of temperature and salinity, the optimum range of temperature and salinity were deflated,there were 18°C-22°C and 30-34.The experiment of nutrition show that:the need of glutamic acid during the development of embryo was more and more.the optimum concentration of glutamic acid was from 0. 5μmol/L move to 2. 5μmol/L when the embryo development from blastula to trochophore-stage larvae. 40μmol/L glutamine can boost the blastula develop to trochophore-stage larvae rapidly. 10 mmol/L taurine can improve the blastula rate best,but when the embryo development the need of taurine was reduced,when the embryo development to trochophore-stage larvae the optimum concentration of taruine was reduced to 5mmol/L. Glucose was not active to the development of blastula, when the embryo develop to gastrulae the need of glucose was become more and more. 2mmol/Lglucose can best improve the trochophore-stage larvae rate, and reduce the developmentarrested larvae rate and abnormal larvae rate rapidly. 0. 009g/L Vc can active to improve the rate of blastula, and the high concentration of Vc make against the embryo development, when the embryo develop into gastrulae and trochophore the need of Vc was reduced gradually.
This study initially prove up the development of the induced embryo and early larvae, and approximately comprehend the need of circumstance and nutrition'factor, and first report the induced embryo and early larvae had temperature and nutrition compensation. This result may afford a new way to capture the problem of high mortality rate during the ploidy breeding. This is more importance to consummate the tetraploid breeding.
本实验总共设2个环境因子和5个营养因子:温度、盐度、谷氨酸、牛磺酸、谷氨酰胺、葡萄糖、维生素C。通过统计各实验组的囊胚、原肠胚、担轮幼虫、畸形幼虫及滞育幼虫的比例,以及检测担轮幼虫和D形幼虫两个时期的幼虫倍性组成,来揭示四倍体栉孔扇贝胚胎及早期幼虫的一些生理特性。结果显示,从囊胚到担轮幼虫,处理组均以20℃组发育最佳,而进入孵化期后,22℃组的孵化率显著高于其他组,这说明了处理组存在着温度补偿,适当地提高温度可以促进四倍体胚胎的孵化。同时发现诱导后的四倍体胚胎对温度和盐度的变化比较敏感,最适盐度和温度范围均缩小了,分别为18-22℃和盐度30-34。而从添加营养因子的实验结果来看:胚胎发育过程中对谷氨酸需求量越来越大,谷氨酸的最佳添加量由囊胚时期的0.5μmol/L上升至孵化时期的2.5μmol/L;在囊胚至担轮幼虫的发育过程中40μmol/L浓度的谷氨酰胺可以显著促进胚胎的发育;10 mmol/L的牛磺酸则可以显著提高囊胚率,但是随着胚胎的进一步发育,其对牛磺酸的需求越来越小,至担轮幼虫时期只有5mmol/L浓度的牛磺酸起作用积极的作用;葡萄糖对囊胚的发育没有积极的帮助,而进入原肠胚时期时,胚胎对葡萄糖浓度的需量逐渐增加,到了担轮幼虫时期,2mmol/L浓度的葡萄糖可以大大提高担轮幼虫的比例,显著降低胚胎的畸形率和滞育率;0.009g/L浓度的维生素C显著促进囊胚的发育,而高浓度的维生素C则抑制胚胎的发育,进入原肠胚时,胚胎对维生素C的需求量减少了,当胚胎发育至担轮幼虫时期时,维生素C添加组除了0.003g/L组外,其他处理组均比对照组低。
本研究初步探明四倍体栉孔扇贝胚胎及早期幼虫的发育情况,以及其对环境和营养的需求情况,首次报道了栉孔扇贝四倍体诱导后的胚胎幼虫存在温度补偿和营养补偿。这将为攻克四倍体倍体育种中出现的高死亡率的问题提供解决方案,对贝类四倍体技术的完善具有重要的参考价值。
In this study, we tested tetraploid induction in Chlamys ferreri by inhititing the release of polar body I(PBl)in newly feitilized eggs. To produce tetraploid,fertilized eggs of Chlamys ferreri were treated with Cytochalasin B (CB) to inhibit the release of the PB I .The development and ploidy composition of induced embryo and early larvae were studied in different circumstance and nutrition's factor.
This study was divided into 7 parts, there were: Temperature,Salinity, Glutamic acid,Taurine,GIutamine,Glucose,Vitamine C. By count the change of the blastula rate, gastrulae rate, hatching rate,trochophore-stage larvae rate, abnormal larvae rate, developmentarrested larvae rate, and the ploidy composition of trochophore-stage and D-stage larvae to open out some traits of physiology on embryo and early larvae of tetraploid Chlamys ferreri .the result show that :from the blastula to trochophore-stage larvae ,20°C was the best temperature to develop the embryo in all treated group.But when the trochophore stage larcae become to hatch,the best temperature was ascended to 22°C ,this explain that there had temperature compensation during the development of ploidy embryo and early larvae.The induced embryo and early larvae were susceptivity to the change of temperature and salinity, the optimum range of temperature and salinity were deflated,there were 18°C-22°C and 30-34.The experiment of nutrition show that:the need of glutamic acid during the development of embryo was more and more.the optimum concentration of glutamic acid was from 0. 5μmol/L move to 2. 5μmol/L when the embryo development from blastula to trochophore-stage larvae. 40μmol/L glutamine can boost the blastula develop to trochophore-stage larvae rapidly. 10 mmol/L taurine can improve the blastula rate best,but when the embryo development the need of taurine was reduced,when the embryo development to trochophore-stage larvae the optimum concentration of taruine was reduced to 5mmol/L. Glucose was not active to the development of blastula, when the embryo develop to gastrulae the need of glucose was become more and more. 2mmol/Lglucose can best improve the trochophore-stage larvae rate, and reduce the developmentarrested larvae rate and abnormal larvae rate rapidly. 0. 009g/L Vc can active to improve the rate of blastula, and the high concentration of Vc make against the embryo development, when the embryo develop into gastrulae and trochophore the need of Vc was reduced gradually.
This study initially prove up the development of the induced embryo and early larvae, and approximately comprehend the need of circumstance and nutrition'factor, and first report the induced embryo and early larvae had temperature and nutrition compensation. This result may afford a new way to capture the problem of high mortality rate during the ploidy breeding. This is more importance to consummate the tetraploid breeding.
引文
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