本文選題：中國鹵蟲 + hyou1　； 參考：《遼寧師范大學(xué)》2016年碩士論文

【摘要】：中國鹵蟲(Artemia sinica)廣泛分布于高鹽水體中,遇到環(huán)境脅迫可以產(chǎn)生休眠卵,其胚胎滯育期新陳代謝減慢,包囊可以抵御低氧、高鹽、寒冷等惡劣環(huán)境的影響。缺氧上調(diào)蛋白Hypoxia up-regulated 1,也被稱為Orp150,其屬于熱休克蛋白家族的成員之一。Hypoxia up-regulated 1在低氧條件下的大鼠膠質(zhì)細(xì)胞中首次被發(fā)現(xiàn),積累在缺氧條件下的內(nèi)質(zhì)網(wǎng)內(nèi),被認(rèn)為在蛋白質(zhì)折疊和分泌中發(fā)揮重要作用。本研究采用RACE技術(shù)首次得到了中國鹵蟲Hypoxia up-regulated 1基因的cDNA全長As-hyou1,同時(shí)也對As-hyou1進(jìn)行了生物信息學(xué)分析和分子進(jìn)化等方面的分析。As-hyou1的基因全長為2952bp,As-hyou1基因包含了116bp的5’非編碼區(qū)以及180bp的3’非編碼區(qū),其中2656bp為開放閱讀框結(jié)構(gòu)。As-hyou1基因所編碼的HYOU1蛋白是一含有886個(gè)氨基酸殘基的蛋白,具有典型的PTZ00009結(jié)構(gòu)域,屬于NBD糖激酶HSP70-actin超家族成員。理論分子量100.5KD,等電點(diǎn)為5.34。Signal P4.1檢測到蛋白序列中含信號肽序列,故中國鹵蟲HYOU1為分泌性蛋白。利用原位雜交技術(shù)對As-hyou1的表達(dá)部位進(jìn)行了分析,As-hyou1廣泛分布于胚胎發(fā)育的各時(shí)期。鹵蟲蟲體生長發(fā)育至已分化階段,鹵蟲的頭胸部、腹部、體表、附肢都會檢測到陽性信號。采用熒光實(shí)時(shí)定量PCR,對As-hyou1的表達(dá)模式進(jìn)行了研究,結(jié)果顯示,As-hyou1在鹵蟲胚胎滯育剛剛結(jié)束階段也就是滯育卵重新回到其正常發(fā)育進(jìn)程時(shí)具有較高的表達(dá)(0h),在早期胚胎發(fā)育期逐漸下調(diào)(5h-15h),在蟲體發(fā)育到無節(jié)幼體時(shí)期表現(xiàn)出上升趨勢;在高鹽刺激下As-hyou1的表達(dá)由50‰開始逐步下調(diào);同時(shí)低溫刺激下表達(dá)量呈下降趨勢。研究初步證明,As-hyou1介導(dǎo)調(diào)節(jié)抑制由缺氧引起的細(xì)胞程序性死亡,并通過內(nèi)質(zhì)網(wǎng)促進(jìn)翻譯、轉(zhuǎn)化和處理其他相關(guān)蛋白。本研究對進(jìn)一步闡明As-hyou1在中國鹵蟲滯育解除、早期胚胎發(fā)育所起的作用,及對中國鹵蟲滯育發(fā)生與解除現(xiàn)象的分子機(jī)制的研究具有較為重要的理論意義。
[Abstract]:Chinese Artemia (Artemia sinica) is widely distributed in high salt water. When exposed to environmental stress, dormant eggs can be produced, the metabolism of embryo diapause is slow, and the cyst can resist the effects of hypoxia, high salt, cold and so on. Hypoxia up-regulated 1, also known as Orp150, is a member of the heat shock protein family. Hypoxia up-regulated 1 was first found in hypoxic rat glial cells and accumulated in the endoplasmic reticulum under hypoxia. It is thought to play an important role in protein folding and secretion. In this study, the full-length As-hyou1 cDNA of Chinese Artemia Hypoxia up-regulated 1 gene was obtained by race technique for the first time. Meanwhile, the As-hyou1 was analyzed in bioinformatics and molecular evolution. The gene length of As-hyou1 gene was 2952bpnAs-hyou1 gene containing the 5'of 116bp. The non-coding region and the 3'non-coding region of 180bp, The HYOU1 protein encoded by the open reading frame structure. As-hyou1 gene of 2656bp is a protein containing 886 amino acid residues. It has a typical PTZ00009 domain and belongs to the superfamily of NBD kinase HSP70-actin. The theoretical molecular weight was 100.5 KD, the isoelectric point was 5.34.Signal P4.1, the signal peptide sequence was detected in the protein sequence, so HYOU1 of Artemia chinensis was secreted protein. The expression site of As-hyou1 was analyzed by in situ hybridization technique. As-hyou1 was widely distributed in all stages of embryonic development. The positive signals were detected in the head, chest, abdomen, body surface and appendage of Artemia. The expression pattern of As-hyou1 was studied by real-time fluorescence quantitative PCR. The results showed that As-hyou1 had high expression (0h) at the end of diapause, which was the stage of diapause, and down-regulated (5h-15h) at the early stage of embryo development. The expression of As-hyou1 decreased gradually from 50 鈥，

本文編號：2082164