皱纹盘鲍生长和抗逆相关新基因的初步研究

IOCAS-IR > 海洋环流与波动重点实验室

	皱纹盘鲍生长和抗逆相关新基因的初步研究
其他题名	Preliminary study on new genes related with growth and resilience of Pacific abalone (Haliotis discus hannai Ino)
	袁泽轶
学位类型	博士
	2007-06-18
学位授予单位	中国科学院海洋研究所
学位授予地点	海洋研究所
关键词	皱纹盘鲍胸腺肽β 荧光定量pcr
摘要	在刘晓研究员已构建的皱纹盘鲍全长cDNA文库及克隆随机测序的结果基础上，分析发现了皱纹盘鲍胸腺肽β（Tβ）cDNA全长，该序列全长487bp，生物信息学分析显示其含有一个135bp的开放阅读框（ORF），编码一个含44个氨基酸的蛋白质分子，另外其5'和3'非翻译区长度分别是69bp和283bp，3'端非翻译区具有AATAAA mRNA加尾信号及PolyA寡聚腺苷酸尾巴。采用荧光定量PCR的方法研究皱纹盘鲍Tβ的组织特异性发现，Tβ在各组织中均有分布，但在外套膜中高表达，推测和其组织结构中富含参与免疫分泌的颗粒有关。利用荧光定量PCR检测感染鳗弧菌后皱纹盘鲍腹足肌中Tβ在不同时间点的转录表达发现，Tβ基因表现出具有显著差异的时间依赖性，12h后Tβ基因的表达量明显升高，且在处理后24h达到最高值，然后表达量开始回落，至处理后72h，Tβ表达量基本降至初始值。推测皱纹盘鲍在受到病原菌侵染后Tβ基因表达量上调可能是由于Tβ基因参与了皱纹盘鲍的免疫反应，它可能在皱纹盘鲍的抗逆机制中发挥着重要作用。发现了一个在皱纹盘鲍大小个体中差异表达的长1555bp含有完整3’端的新基因片断，暂命名为TH。利用荧光定量PCR检测皱纹盘鲍RwRh、J1Rh、J1Jm、RwJm 4个家系大小个体中TH基因的差异表达，发现其在大个体中的表达量显著高于小个体，推测其可能与皱纹盘鲍生长速度有某种关联。荧光定量PCR对皱纹盘鲍TH基因时空差异表达的研究发现，其在发育过程中的表达呈现出一定的规律性，即在围口壳幼虫期前始终保持较低水平的表达，当围口壳形成后表达量急剧增加，至幼鲍时达到最高值，在成鲍中也维持较高水平的表达，提示该基因可能调控壳的发育。同时，皱纹盘鲍TH基因具有组织特异性表达，在外套膜中的表达量显著高于其他组织，推测和其组织结构中富含参与壳形成的颗粒有关。另外还比较了皱纹盘鲍RwRh、J1Rh、J1Jm、RwJm 4个家系中TH基因的差异表达，结果发现在这4个家系中的相对表达呈现出一定的规律，即J1Rh＞RwRh＞RwJm＞J1Jm，可能有助于解释家系之间生物学性状上的差异。
其他摘要	Theβ-Thymosin was obtained through sequencing clones picked randomly in the cDNA library constructed from Haliotis discus hannai Ino which was accomplished by Liu Xiao, suggesting some full-length cDNAs were contained in the library. Tβ was 487 bp in length, including an open reading frame of 135 bp, which contained a deduced amino acid sequence of 44 residues. It consisted of a 5'-terminal untranslated region (UTR) of 69bp, a 3'-terminal UTR of 283bp with a canonical polyadenylation singnal sequence AATAAA and a Poly (A) tail. The expression of Tβ mRNA of Haliotis discus hannai Ino in the different tissues was analyzed by real-time PCR. It was detected in all tissues and expressed highly in the mantle. We speculate that it owes to grains related to immunity in this tissue. And the temporal expression of Tβwas measured by quantitative RT-PCR after bacterial challenge. The mRNA levels in muscle varied at different infection time: At 12h post-infection, it increased dramatically, the highest was at 24h post-infection, followed by a recovery to normal level after 72h. The up-regulated mRNA expression of Tβ in Haliotis discus hannai Ino following infection response indicates that the Tβgene is inducible and involved in immune response and it may also play an important role in Haliotis discus hannai Ino immune mechanism. A new gene expressed differently between big and small individuals of Pacific abalone was obtained. It was named TH provisionally, consisting of a 3'-terminal region with 1555 bp in length. The expression of TH of Haliotis discus hannai Ino between big and small individuals in the 4 families (RwRh、J1Rh、J1Jm、RwJm)was analyzed by real-time PCR. It expressed more highly in big individuals. We speculated that it may relate to the rate of growth of Pacific abalone. Spatial and temporal expression pattern during Pacific abalone development of TH gene was investigated using real-time PCR. It presented some rules during development. Before creeping larva stage, it always expressed at low level. After creeping larva stage, the expression rose sharply and highest at the infant estate. And it kept high level during adult. It was likely to regulate the development of shell. The TH was expressed differently in tissues of Pacific abalone. It was detected expressed highly in the mantle. We speculate that it owes to grains related to forming of shell in this tissue. In addition, the relative expression of TH in the 4 families (RwRh、J1Rh、J1Jm、RwJm)was analyzed by real-time PCR. The result was that: J1Rh＞RwRh＞RwJm＞J1Jm. It might be helpful to explain the different characters among families.
页数	74
语种	中文
文献类型	学位论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/795
专题	海洋环流与波动重点实验室
推荐引用方式 GB/T 7714	袁泽轶. 皱纹盘鲍生长和抗逆相关新基因的初步研究[D]. 海洋研究所. 中国科学院海洋研究所,2007.

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