家蚕卵形指数对孵化率的影响及孵化相关基因表达分析

Effects of egg shape index of silkworm(Bombyx mori)on hatching rate and related gene expression analysis

[目的]分析家蚕卵形指数对孵化率的影响规律,以及孵化酶基因(BmHEⅠ和BmHEⅡ)、几丁质酶基因(Chitinase)和周期蛋白基因(Period)在不同卵形间的表达规律,为阐明家蚕卵形指数与孵化率的相关性及其分子机理打下基础。[方法]调查家蚕Nistari品种野生型正常形品种(Nistari+)及其新突变体纺锤形品种(Nistari-)的卵形指数、孵化率、受精率和转青死卵率,分析其卵形指数与孵化率的相关性;并通过实时荧光定量PCR检测蚕卵胚胎发育后期孵化相关基因的表达情况。[结果lNistari+蚕卵的平均卵形指数为1.26,极显著低于Nistari-蚕卵(1.73)(P<0.01,下同);孵化率为96.22%,极显著高于Nistari-蚕卵(51.33%)。当Nistari+蚕卵的卵形指数为1.24~1.25时,其孵化率(97.39%)和受精率(98.84%)均最高,转青死卵率最低(2.61%);而Nistari-蚕卵的卵形指数为1.77~1.81时,其孵化率(56.52%)和受精率(74.87%)均最高,转青死卵率最低(43.48%)。相关分析结果表明,Nistari蚕卵孵化率、受精率和转青死卵率与卵形指数存在一定的相关性,且受卵形指数影响。在蚕卵胚胎发育后期(孵化前),Nistari+蚕卵的BmHEⅠ、BmHEⅡ、Chitinase和Period基因大量表达,对应的相对表达量显著(P<0.05)或极显著高于NISTAR-蚕卵。[结论]家蚕Nistari品种的卵形指数与孵化率存在一定相关性,而BmHEⅠ、BmHEⅡ、Chitinase和Period基因的表达差异是导致Nistari-蚕卵与Nistari+蚕卵孵化率差异的分子基础。

[Obj ective]The effects of silkworm egg shape index(ESI) on hatching rate(HR),and the expression patterns of hatching enzyme genes(BmHE Ⅰ and BmHE Ⅱ),chitinase gene(Chitinase) and period protein gene(Period) in different egg shapes were investigated,which laid a foundation for elucidating the correlation between egg shape index and hatching rate of silkworm eggs and its molecular mechanism.[Method] The ESI,HR,fertilization rate(FR),and mortality rate in egg with pigmentary body of embryo(MR) were investigated between the wild type normal egg variety(Nistari+)and its new mutant spindle egg variety(Nistari),and the correlation of ESI with HR were analyzed.And investigated the differences in gene expression levels related to hatching process in the later stages of embryonic development by real-time quantitative PCR.[Result]The mean ESI of Nistari+was 1.26,which was extremely lower than that of Nistari-(1.73)(P<0.01,the same below);the HR of Nistari+was 96.22%,which was extremely higher than that of Nistari-silkworm eggs(51.3 3 %).When the ESI of Nistari+was 1.24-1.25,the HR(97.39%) and FR(98.84%) were the highest and the MR(2.61%) was the lowest,while when the ESI of Nistari-was 1.77-1.81,the HR(56.52%) and FR(74.87%) of its eggs were the highest,MR was the lowest(43.48%).The results of correlation analysis showed that there was a certain correlation between the Nistari ESI and HR,FR and MR,and was affected by ESI.In the later stages of silkworm egg embryo development(before hatching),the expression levels of BmHE Ⅰ,BmHE Ⅱ,Chitinase and Period genes of Nistari+silkworm eggs were high,and were significantly(P<0.05) or extremely(P<0.01) higher than those of Nistari-silkworm eggs(P<0.05).[Conclusion] There is certain correlation between ESI and HR of silkworm Nistari varieties.The differences in expression of four genes,BmHE Ⅰ,BmHE Ⅱ,Chitinase and Period,are the molecular basis for the differences in HR between silkworm varieties of Nistari-and Nistari+.