22 resultados para Melanogaster

em Chinese Academy of Sciences Institutional Repositories Grid Portal


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Nucleotide sequences of the spacer region of the histone gene H2A-H2B from 36 species of Drosophila melanogaster species group were determined. The phylogenetic trees were reconstructed with maximum parsimony, maximum likelihood, and Bayesian methods by u

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A central goal of evolutionary genetics is an understanding of the forces responsible for the observed variation, both within and between species. Theoretical and empirical work have demonstrated that genetic recombination contributes to this variation by breaking down linkage between nucleotide sites, thus allowing them to behave independently and for selective forces to act efficiently on them. The Drosophila fourth chromosome, which is believed to experience no-or very low-rates of recombination has been an important model for investigating these effects. Despite previous efforts, central questions regarding the extent of recombination and the predominant modes of selection acting on it remain open. In order to more comprehensively test hypotheses regarding recombination and its potential influence on selection along the fourth chromosome, we have resequenced regions from most of its genes from Drosophila melanogaster, D. simulans, and D. yakuba. These data, along with available outgroup sequence, demonstrate that recombination is low but significantly greater than zero for the three species. Despite there being recombination, there is strong evidence that its frequency is low enough to have rendered selection relatively inefficient. The signatures of relaxed constraint can be detected at both the level of polymorphism and divergence.

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Intron loss and its evolutionary significance have been noted in Drosophila. The current study provides another example of intron loss within a single-copy Dfak gene in Drosophila. By using polymerase chain reaction (PCR), we amplified about 1.3 kb fragment spanning intron 5-10, located in the position of Tyr kinase (TyK) domain of Dfak gene from Drosophila melanogaster species group, and observed size difference among the amplified DNA fragments from different species. Further sequencing analysis revealed that D. melanogaster and D. simulans deleted an about 60 bp of DNA fragment relative to other 7 Drosophila species, such as D. elegans, D. ficusphila, D. biarmipes, D. takahashii, D. jambulina, D. prostipennis and D. pseudoobscura, and the deleted fragment located precisely in the position of one intron. The data suggested that intron loss might have occurred in the Dfak gene evolutionary process of D. melanogaster and D. simulans of Drosophila melanogaster species group. In addition, the constructed phylogenetic tree based on the Dfak TyK domains clearly revealed the evolutionary relationships between subgroups of Drosophila melanogaster species group, and the intron loss identified from D. melanogaster and D. simulans provides a unique diagnostic tool for taxonomic classification of the melanogaster subgroup from other group of genus Drosophila.

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Polysaccharides isolated from Porphyra (porphyran) have been known to have diverse biological activities, including immunomodulatory and antioxidant activities. The molecular weight-antiaging activity relationship of degraded porphyrans was examined in this study. Natural porphyran was extracted from P. haitanensis, and then was degraded into different molecular weight fractions, P1 molecular weight 49 kDa, P2 molecular weight 30 kDa, P3 molecular weight 8.2 kDa, by free radical. The influence on life span and vitality of porphyrans were carried out on Drosophila melanogaster. We found that all the degraded porphyrans and natural porphyran (P), added daily to the diet, can significantly increase the life span of D. melanogaster, except for P3. Among them, P1 exhibited the most prolonging life span activity. Furthermore, vitality of middle-aged flies (assessed by measuring their mating capacity) receiving porphyrans was increased considerably in comparison with the controls. Finally, in the heat-stress test, we observed a remarkable increase in survival time, especially in P3-diet groups. These results suggest that porphyrans may be effective in reducing the rate of the aging process and molecular weight has important influence on the effects. It seems that P1 and P2, possessed higher molecular weight, may be more useful in normal metabolic condition and P3, possessed the lowest molecular weight, may be more beneficial for D. melanogaster in stress condition. (C) 2007 Elsevier Ltd. All rights reserved.

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Regulation of neuronal gene expression is critical to nervous system development. REST (RE1-silencing transcription factor) regulates neuronal gene expression through interacting with a group of corepressor proteins including REST corepressors (RCOR). Here we show that Xenopus RCOR2 is predominantly expressed in the developing nervous system. Through a yeast two-hybrid screen, we isolated Xenopus ZMYND8 (Zinc finger and MYND domain containing 8) as an XRCOR2 interacting factor. XRCOR2 and XZMYND8 bind each other in co-immunoprecipitation assays and both of them can function as transcriptional repressors. XZMYND8 is co-expressed with XRCOR2 in the nervous system and overexpression of XZMYND8 inhibits neural differentiation in Xenopus embryos. These data reveal a RCOR2/ZMYND8 complex which might be involved in the regulation of neural differentiation. (C) 2010 Elsevier Inc. All rights reserved.

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Drosophila (Sophophora) subpulchrella Takamori and Watabe, sp. nov., of the D. suzukii subgroup in the D. melanogaster species group, is described from Japan and southern China, and compared with its sibling species, D. pulchrella Tan et al. distributed i

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The fourth chromosome of Drosophila melanogaster and its sister species are believed to be nonrecombining and have been a model system for testing predictions of the effects of selection on linked, neutral variation. We recently examined nucleotide variat

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The genus Pseudostegana is revised, with descriptions of 20 new species from Southeast Asia: P. angustifasciata Chen and Wang, sp. n., P. atrofrons Chen and Toda, sp. n., P. bifasciata Chen and Toda, sp. n., P. bisetosa Chen and Toda, sp. n., P. curvata Chen and Toda, sp. n., P. dactylis Chen and Toda, sp. n., P. dolichopoda Chen and Wang, sp. n., P. hamata Chen and Toda, sp. n., P. latifasciata Chen and Toda, sp. n., P. leptoptera Chen and Toda, sp. n., P. melanogaster Chen and Toda, sp. n., P. melanopogonias Chen and Toda, sp. n., P. myrmecoformis Chen and Toda, sp. n., P. nitidifrons Chen and Wang, sp. n., P. nitidiventris Chen and Toda, sp. n., P. orbicapitata Chen and Toda, sp. n., P. oxycephala Chen and Toda, sp. n., P. pallidimaculata Chen and Wang, sp. n., P. philoga Chen and Wang, sp. n. and P xanthoptera Chen and Wang, sp. n. On the basis of the wing patterns, six species-groups are established: the atrofrons, grandipalpis, fleximediata, javana, latiparma and zonaria groups. A key to the all species of this genus is provided.

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本论文结合功能研究和进化遗传学方法对动物天然免疫(innate immunity)相关分子的进化历程进行深入研究。受体对病原微生物的识别是天然免疫系统发挥功能的基础。作为模式识别受体(pattern recognition receptor, PRR),果蝇肽聚糖识别蛋白SD(PGRP-SD)在识别革兰氏阳性细菌的过程中发挥了重要作用。针对已有的黑腹果蝇(Drosophila melanogaster)群体数据,我们发现PGRP-SD在群体中存在2类高频的等位基因(分别为等位基因1和等位基因2)。以D. simulans为外群,我们追溯了黑腹果蝇2类等位基因上氨基酸的变化。这些氨基酸的结构特征和在蛋白质上所处的位置提示这2类等位基因在功能方面可能存在分化。通过功能研究的方法,我们发现在黑腹果蝇中该基因功能方面发生了显著的变化。等位基因2在有微生物时能激活天然免疫反应,但等位基因1的转基因果蝇成虫只要有外伤即便没有微生物的情况下即能激发天然免疫反应,而带有等位基因2果蝇成虫则不具有该功能。这一结果提示我们,发生在该等位基因上的氨基酸变化导致了其识别功能的变化。与推导的祖先基因相比,等位基因1发生了一个氨基酸的变化,因此导致其功能从识别细菌细胞壁组分肽聚糖转变为一未知的自身组分,即从病原相关分子模式(pathogen-associated molecular pattern,PAMP)识别受体转变为损伤相关识别模式(damage-associated molecular pattern, DAMP)识别受体。通过这一功能变化, 果蝇成虫可以通过仅识别自身损伤即可激活相应的免疫反应,对后续可能侵入的微生物进行杀伤。已有研究结果显示,微生物在进化过程中已经形成针对DAMP和PAMP规避策略。上述2类等位基因的同时存在能使黑腹果蝇同时具备两个机制,更加充分地抵抗病原微生物的入侵。结合功能研究和针对自然群体的群体遗传学分析,我们认为在黑腹果蝇群体中以高频共存的2类PGRP-SD等位基因可能可能受到了平衡选择(balancing selection)作用。上述工作主要研究了天然免疫系统识别受体的进化。而本论文的另一部分则主要针对天然免疫系统的效应分子(effector)进行了研究。作为重要的效应分子,抗菌肽在杀菌方面发挥着最为直接的作用。因此,研究抗菌肽的进化对于探索天然免疫系统的进化具有重要意义。本研究以两栖类动物大蹼铃蟾抗菌肽基因家族为例,通过对分别来自2个大蹼铃蟾个体的皮肤cDNA文库进行测序,我们鉴别出56个不同的抗菌肽cDNA序列。每一个cDNA均编码2个不同的抗菌肽,maximin 和maximin H。基于针对这些cDNA序列的分析,我们发现2类抗菌肽编码序列的非同义替代率均高于同义替代率,呈现高度分化的特征。但是,在信号肽和其它非抗菌肽编码区域并没有发现这种情况。这一结果提示抗菌肽可能受到超显性选择(overdominent selection, 即平衡选择)的影响。同时,我们分别从皮肤和肝脏克隆基因了7个抗菌肽的基因组编码序列并进行了测序。这些从不同组织获得的抗菌肽在各个编码序列中均存在序列的差异的同时呈现了相同的结构。这一结果提示不同抗菌肽间的差异不太可能来自于体细胞突变而是快速序列进化的结果。通过构建来自于同一个体的抗菌肽的不同编码区的基因树,我们发现结构域重排(domain shuffling)和/或基因转换(gene conversion)在这些抗菌肽的进化历程中发挥作用。

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本实验室果蝇研究工作,主要集中在黑腹果蝇的新基因起源的研究。新基因起源的分子机制主要包括:外显子重排、基因复制、基因逆转座、移动元件介导、基因水平转移、基因从头起源、基因的断裂融合。为了阐述这些新基因的产生和它们所带来的物种适应性,我们对这些新近起源的基因进行了功能研究。但是,仅仅限于新基因所在物种的功能研究并不能完全解释新基因产生的进化原因,我们需要了解它是否能够给没有该基因的果蝇物种带来一定的适应性。例如一些生殖相关新基因,如果我们将它们转入没有该基因的果蝇,那是否能够给该果蝇带来生殖能力的提高?无论结果如何,这都为我们研究新基因的起源提供一个重要线索。由此,黑腹果蝇以外的其它果蝇物种中实现转基因成为该研究的重要技术环节。但是,实验室目前的转基因系统仅限于P转座子介导的黑腹果蝇转基因系统,因而我们需要建立一种新的转基因平台。而转座子Minos打破物种范围的转基因特性,以及它的转座特点为我们提供了选择。转座子Minos是从果蝇D. hydei中克隆出来长约1.8kb的Ⅱ型转座子,Tc1家族转座元件成员。Minos的转座机制与大部分转座子一样,在宿主基因组里面实行着剪切和粘贴的运作机制。Minos在转座时,偏向插入TA位点并且主要集中于内含子区域,这样可以减少对插入位置基因的影响。此外,Minos在黑腹果蝇中的转座效率约30%,并且拥有一套成熟的选择标记。因此,Minos成为我们解决非黑腹果蝇转基因技术难题的首选。 在本文的工作中,我们采用由希腊Savakis教授(希腊分子生物学与生物技术研究所)提供的Minos转基因系统,完成果蝇的转基因实验。在这套转基因系统中,非自主的转座子Minos和转座酶基因被克隆到了不同载体当中。其中Minos转座子序列中插入了由3xP3眼睛特异表达的启动子介导表达的eGFP报告基因,而转座酶基因则由热激蛋白hsp70启动子调控表达。实验过程中,我们在果蝇D. melanogaster 和D. yakuba的胚胎中分别同时显微注射入含有转座子和转座酶本实验室果蝇研究工作,主要集中在黑腹果蝇的新基因起源的研究。新基因起源的分子机制主要包括:外显子重排、基因复制、基因逆转座、移动元件介导、基因水平转移、基因从头起源、基因的断裂融合。为了阐述这些新基因的产生和它们所带来的物种适应性,我们对这些新近起源的基因进行了功能研究。但是,仅仅限于新基因所在物种的功能研究并不能完全解释新基因产生的进化原因,我们需要了解它是否能够给没有该基因的果蝇物种带来一定的适应性。例如一些生殖相关新基因,如果我们将它们转入没有该基因的果蝇,那是否能够给该果蝇带来生殖能力的提高?无论结果如何,这都为我们研究新基因的起源提供一个重要线索。由此,黑腹果蝇以外的其它果蝇物种中实现转基因成为该研究的重要技术环节。但是,实验室目前的转基因系统仅限于P转座子介导的黑腹果蝇转基因系统,因而我们需要建立一种新的转基因平台。而转座子Minos打破物种范围的转基因特性,以及它的转座特点为我们提供了选择。转座子Minos是从果蝇D. hydei中克隆出来长约1.8kb的Ⅱ型转座子,Tc1家族转座元件成员。Minos的转座机制与大部分转座子一样,在宿主基因组里面实行着剪切和粘贴的运作机制。Minos在转座时,偏向插入TA位点并且主要集中于内含子区域,这样可以减少对插入位置基因的影响。此外,Minos在黑腹果蝇中的转座效率约30%,并且拥有一套成熟的选择标记。因此,Minos成为我们解决非黑腹果蝇转基因技术难题的首选。 在本文的工作中,我们采用由希腊Savakis教授(希腊分子生物学与生物技术研究所)提供的Minos转基因系统,完成果蝇的转基因实验。在这套转基因系统中,非自主的转座子Minos和转座酶基因被克隆到了不同载体当中。其中Minos转座子序列中插入了由3xP3眼睛特异表达的启动子介导表达的eGFP报告基因,而转座酶基因则由热激蛋白hsp70启动子调控表达。实验过程中,我们在果蝇D. melanogaster 和D. yakuba的胚胎中分别同时显微注射入含有转座子和转座酶所在的质粒。转座酶在37度条件诱导下进行表达,协助Minos完成转座过程。在转基因果蝇的阳性筛选中,我们利用眼睛特异表达的绿色荧光蛋作为选择标记。并且,我们通过PCR实验进一步验证了转基因果蝇的真实性。本研究中,我们对转基因实验条件进行了初步优化。我们通过对黑腹果蝇白眼突变品系W1118和D. yakuba注射后胚胎进行保湿,对D. yakuba注射胚胎进行非退壳处理。在改进条件下W1118和D. yakuba的存活率分别为10%和3%左右。通过筛选转基因阳性果蝇,我们得出Minos在W1118和D. yakuba中的转座效率分别在32%和20%左右。我们的实验结果再一次证实了Minos在果蝇D. melanogaster中可行性。同时,该工作也初步完成了在果蝇D. yakuba 中的第一次Minos介导的转基因实验,为新基因的跨物种功能研究奠定了实验基础。在未来的工作计划中,我们将采用Minos转基因系统,把实验室目前研究的黑腹果蝇新基因导入其它物种果蝇进行功能研究。 水稻是一种重要的世界粮食作物,世界上过半的人口以水稻为主食。水稻相对别的粮食作物来讲具有较小的基因组,并且拥有较好的基因组注释,是一种理想的单子叶模式生物。植物转基因技术的发展推动着水稻功能基因组学的研究,目前水稻的转基因技术主要依赖于土壤细菌农杆菌(Agrobacterium tumefaciens)T-DNA介导的外源基因染色体插入。在自然状态下,农杆菌的T-DNA位于Ti致瘤质粒当中。它包括了一些转座元件和一些帮助T-DNA转座的毒性蛋白基因和调节基因。由于Ti质粒上的T-DNA太长,并且没有太多的酶切位点,因此自然状态的T-DNA不适合进行转基因实验。为了方便T-DNA的实际应用,研究人员创立了双载体转基因系统。T-DNA转座区被分离到出Ti载体,并且装载到另外一个适合实验操作的质粒当中,而毒性蛋白表达基因等则保留在Ti质粒上。因此,在进行T-DNA介导的转基因实验时,需要同时存在T-DNA载体和Ti质粒。 本文以“水稻注释计划数据库RAP-DB”的表达数据为参考,选择了60个高表达基因的启动子区域进行克隆。通过对T-DNA载体pCAMBIA1301 进行改造,去掉其原来的35S启动子,将预测的基因启动子克隆到该载体中并与报告基 摘要 因GUS 基因融合。通过分子克隆实验,我们得到了45个高表达基因的启动子载体。最终,为了测试这45个启动子的启动效率,我们会将它们转化到水稻愈伤组织中通过启动子融合的GUS基于表达情况来判断我们启动子的启动效率。

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双尾-C 基因 (Bicaudal-C)首先在果蝇(Drosophila melanogaster)中发现,其功能丧失导致果蝇胚胎滤泡细胞的错误迁移、头部的缺失和双尾结构的形成。后来发现多个物种都含有Bicaudal-C 的同源基因,其中小鼠中的同源基因Bicc1 的缺失导致小鼠产生肾脏等脏器的病变,其症状与人类多囊肾疾病高度相似,但其具体机制还不清楚。本研究以小鼠肾脏组织总RNA 为模板体外反转录为cDNA,通过分段巢式 PCR 及酶切连接的方法获得了全长约3Kb 的小鼠Bicc1 cDNA 序列。根据生物信息学分析全长的Bicc1 蛋白,选择两个免疫原性较好的区段作为抗原位点构建相应的原核表达载体;IPTG 诱导表达并纯化融合蛋白,制备两种兔抗Bicc1 蛋白多克隆抗体,并通过Western blot 证实这两种抗体具有高度特异性。用细胞免疫荧光方法及免疫组织化学方法对该蛋白的定位做了一些初步研究。发现Bicc1 蛋白定位于体外培养的小鼠肾细胞的细胞质内,并在胚胎发育于期表达仅在心脏,后来逐步地在各个组织器官内出现,并在出生后的小鼠体内表达稳定。Bicc1 mDNA 也表达于多个器官内,并且在肾脏中有明显较高的表达量。找到了的两个针对Bicc1 基因的RNAi 的序列,通过荧光强度变化和Western blot 均证明这两个序列能明显降低Bicc1 蛋白在体外培养细胞中的表达水平,为下一步建立稳定的细胞株奠定了良好的基础。

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Tumor necrosis factor receptors (TNFRs) are a superfamily of proteins characterized by the unique cysteine-rich domain (CRD) and their important roles in diverse physiological and pathological events such as inflammation, apoptosis, autoimmunity and organogenesis. The first member of the molluscan TNFR family, designated as CfTNFR, was identified from Zhikong scallop Chlamys farreri by expressed sequence tag (EST) and rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of CfTNFR was of 1334 bp, consisting of a 5' UTR of 17 bp, a 3'UTR of 69 by with a poly (A) tail, and an open reading frame (ORE) of 1248 by encoding a polypeptide of 415 amino acids with a theoretical isoelectric point of 8.33 and predicted molecular weight of 47.07 kDa. There were a signal peptide, a CRD, a transmembrane region and a death domain in the deduced amino acid sequence of CfTNFR, suggesting that it was a typical type 1 membrane protein. The high identities (22-40%) of CfTNFR with other TNFR superfamily members indicated that CfTNFR should be a member of TNFR superfamily, and moreover, it should be the first death domain-containing TNFR found in invertebrates. Phylogenetic analysis revealed that CfTNFR was closely related to TNFR-like proteins from Strongylocentrotus purpuratus, Drosophila melanogaster and Ciona intestinalis, and they formed a separate branch apart from vertebrate TNFRs. The spatial expression of CfTNFR transcripts in healthy and bacteria challenged scallops was examined by quantitative real-time PCR. CfTNFR transcripts could be detected in all tested tissues, including haemocytes, gonad, gill, mantle and hepatopancreas, and significantly up-regulated in the tissues of gonad, gill, mantle and hepatopancreas after Listonella anguillarum challenge, indicating that CfTNFR was constitutive and inducible acute-phase protein involved in immune defence. The present results suggested the existence of the TNFR-like molecules and TNF-TNFR system in low invertebrates, and provided new insights into the role of CfTNFR in scallop innate immune responses to invading microorganisms. (C) 2009 Elsevier Ltd. All rights reserved.

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GRP78 (78 kDa glucose-regulated protein), also known as BiP (immunoglobulin heavy-chain-binding protein), is an essential regulator of endoplasmic reticulum (ER) homeostasis because of its multiple functions in protein folding, ER calcium binding, and controlling of the activation of transmembrane ER stress sensors. In this report, we cloned the full length cDNA of GRP78 (FcGRP78) from Chinese shrimp Fenneropenaeus chinensis. This cDNA revealed a 2,325 bp with 1,968 bp open reading frame encoding 655 amino acids. This is the first reported GRP78 gene in Crustacea. The deduced amino acid sequence of FcGRP78 shared high identity with previously reported insect GRP78s: 86, 87 and 85% identity with GRP78s of Drosophila melanogaster, Aedes aegypti and Bombyx mori, respectively. Northern blot analysis shows that FcGRP78 is ubiquitously expressed in tissues of shrimp. Heat shock at 35A degrees C significantly enhanced the expression of FcGRP78 at the first hour, reached the maximum at 4 h post heat shock, dropped after that and resumed to the normal level until 48 h of post recovery at 25A degrees C. Additionally, differential expression of FcGRP78 was detected in haemocytes, hepatopancreas and lymphoid organ when shrimp were challenged by white spot syndrome virus (WSSV). We inferred that FcGRP78 may play important roles in chaperoning, protein folding and immune function of shrimp.

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Previously we suggested that four proteins including aldolase and triose phosphate isomerase (TPI) evolved with approximately constant rates over long periods covering the whole animal phyla. The constant rates of aldolase and TPI evolution were reexamined based on three different models for estimating evolutionary distances, It was shown that the evolutionary rates remain essentially unchanged in comparisons not only between different classes of vertebrates but also between vertebrates and arthropods and even between animals and plants, irrespective of the models used, Thus these enzymes might be useful molecular clocks for inferring divergence times of animal phyla, To know the divergence time of Parazoa and Eumetazoa and that of Cephalochordata and Vertebrata, the aldolase cDNAs from Ephydatia fluviatilis, a freshwater sponge, and the TPI cDNAs from Ephydatia fluviatilis and Branchiostoma belcheri an amphioxus, have been cloned and sequenced, Comparisons of the deduced amino acid sequences of aldolase and TPI from the freshwater sponge with known sequences revealed that the Parazoa-Eumetazoa split occurred about 940 million years ago (Ma) as determined by the average of two proteins and three models, Similarly, the aldolase and TPI clocks suggest that vertebrates and amphioxus last shared a common ancestor around 700 Ma and they possibly diverged shortly after the divergence of deuterostomes and protostomes.

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Calreticulin (CRT), as an endoplasmic reticulum luminal resident protein, plays important roles in Ca2+ homeostasis and molecular chaperoning. CRT on the surface of the cell can modulate cell adhesion, phagocytosis and integrin-dependent Ca2+ signaling. The full length cDNA of calreticulin (FcCRT) was cloned from Chinese shrimp Fenneropenaeus chinensis. It consists of 1672 by with an open reading frame of 1221 bp, encoding 406 amino acids. This is the first reported cDNA sequence of calreticulin in Crustacea. The deduced amino acid sequence of FcCRT showed high identity with those of Bombyx mori (88%), Drosophila melanogaster (83%), Mus musculus (82%) and Homo sapiens (82%). Highest expression of FcCRT was detected in ovary by Northern blot and in situ hybridization. Different mRNA levels of FcCRT were detected at various molting stages. Expression of FcCRT was induced significantly after 3 h of heat shock treatment, reached the maximum at 4 h and dropped after that. Differential expression profiles of FcCRT were observed in hepatopancreas and haemocytes when shrimp were challenged by white spot syndrome virus (WSSV). From the above results, we inferred that FcCRT might play important roles in Ca2+ homeostasis, chaperoning and immune function in shrimp. (c) 2007 Elsevier Inc. All rights reserved.