双斑东方鲀染色体核型分析

鱼类的染色体核型分析是鱼类遗传学研究的基础,作为我国重要养殖品种的双斑东方鲀(Takifugu bimaculatus)尚未发现有染色体核型的相关报道.采用体内注射植物血球凝集素(PHA)及秋水仙素法制备了双斑东方鲀头肾组织染色体标本并分析其核型.结果表明,双斑东方鲀二倍体染色体数为2n=44,其染色体臂数为NF=62,核型公式为12m+6sm+26t.通过比较分析推测2n=44应该是东方鲀属的染色体基础核型,而其染色体核型的高度保守可能是东方鲀属鱼类易于发生种间杂交的主要原因之一.厦门南方海洋研究中心资助项目(13GYY002NF07);;海洋经济创新发展区域示范专项资助项目(12PYY001SF08-XMDX-2

福建石斑鱼产业发展形势分析

石斑鱼是重要的海水养殖鱼类,福建省的石斑鱼全产业链产值超50亿元,2017年养殖产量占全国总产量的22.10%。笔者在石斑鱼产业发展调研过程中发现石斑鱼产业仍面临育苗水平不高,养殖病害较严重的挑战,建议实施种业创新、建设示范基地、拓展精深加工及培育特色品牌,推进产业转型升级

Genome size for six commercially important fishes in China

采用流式细胞术,以人(HOMO SAPIEnS)淋巴细胞dnA含量(7.00Pg/2C)为标准,以鱼的红血细胞为材料,测定中国6种重要经济鱼类条纹斑竹鲨(CHIlOSCyllIuM PlAgIOSuM)、斜带石斑鱼(EPInEPHEluS COIOIdES)、赤点石斑鱼(EPInEPHEluS AkAArA)、大黄鱼(PSEudOSCIAEnA CrOCEA)、中华乌塘鳢(bOSTrICHTHyS SInEnSIS)、大菱鲆(SCOPHTHAlMuS MAXIMuS)的基因组大小(C-值)。结果显示,这6种经济鱼类的单倍体dnA含量分别为:条纹斑竹鲨(4.91±0.24)Pg;斜带石斑鱼(1.25±0.04)Pg;赤点石斑鱼(1.23±0.11)Pg;大黄鱼(0.76±0.03)Pg;中华乌塘鳢(0.85±0.04)Pg;大菱鲆(0.65±0.01)Pg。研究结果可为这6种经济鱼类的种质鉴定提供依据。此外,根据本研究的实验结果并结合其他鱼类的基因组大小资料,对鱼类基因组大小与其染色体数目及进化地位之间的相关性进行了探讨,对比分析显示,鱼类进化地位越高则dnA含量越少,同时鱼类基因组的大小差异存在复杂性,并不完全与染色体数目相关。本研究还就鱼类基因组大小的测定方法等问题提出建议,认为应采用流式细胞术和人淋巴细胞作为规范的检测方法和对照标准。The study of the genome size is highly significant as it can not only help to instruct the hybridization and breeding program,but also aid to the whole-genome sequencing project of commercial fishes.Chiloscyllium plagiosum,Epinephelus coioides,E.akaara,Pseudosciaena crocea,Bostrichthys sinensis and Scophthalmus maximus,with high market value,are commercially important and popular fishes in China because of their delicious and nutritious meat.In China,E.coioides,P.crocea and S.maximus have been the key of the aquaculture industry over a large scale.Nevertheless,there are still limited data of the genome size despite of many corresponding researches on the heredity,evolution and breeding.In this paper,the genome size( C-value) of these fishes was determined using a flow cytometry technique,and Human( Homo sapiens)lymphocyte( 3.50 pg/C) was serving as an internal control.The results showed C-values are 4.91±0.24,1.25±0.04,1.23±0.11,0.76±0.03,0.85±0.04,and 0.65±0.01 for C.plagiosum,E.coioides,E.akaara,P.crocea,B.sinensis and S.maximus,respectively.The results showed that the genome size of the class Chondrichthyes was significantly larger than that of Osteichthyes,and the genome size was found significantly different among these fishes except for E.coioides and E.akaara,which are in the same genus and have an affinity relationship between them.The C-values may be used as a criterion for species identification among these fishes.Base on our results and other data from genome-size database which were measured using the same method and control,the relationship between C-values and evolution status and chromosome amount of the fishes was discussed.Most of the data showed that with the higher evolution status of the fish,the lower C-value is presented.To a certain extent,the result demonstrated that C-value prefers to be more condensed along with the evolution of the genome.However,there are many complex factors which can influence the genome size of the fish.The point of view that the genome size positively correlates to the amount of chromosome is not always correct according to the data.In the present study,the amount of chromosome of P.crocea and B.sinensis are 48 and 46 respectively,while their relative genome sizes are 0.76 pg and 0.85 pg.Furthermore,the result also shows that there is more diversity on the genome size among species existing in original order such as the Chondrichthyes and Acipenseriformes of the Osteichthyes,which may be due to an age long evolution history of this fish.Finally,in order to avoid the disadvantages of poor accuracy and low comparability,we consider that the use of the flow cytometry technique and human lymphocyte as a control should be a more appropriate criterion for genome size determination protocol.国家自然科学基金项目(40976094

Molecular phylogenetic relationships of 30 grouper species in China Seas based on 16S rDNA fragment sequences

石斑鱼因其种类繁多、分布广泛及缺乏显著的形体特征,使其系统分类的研究颇为困难。为探讨中国近海石斑鱼类的系统进化关系,通过PCR扩增获得了石斑鱼亚科(Epinephelinae)6属30个种类的线粒体16SrDNA基因片段序列。采用多个生物软件对序列变异和碱基组成进行分析,计算了Kimura2parameter遗传距离、转换/颠换比等遗传信息指数,并结合GenBank石斑鱼属的同源序列,以多纹长尾(Pronotogrammusmultifasciatus)和皮氏叫姑鱼(Johniusbelengerii)为外群构建NJ、MP和ML系统树。根据所得分子依据并结合形态学特征,推论如下:1)在本研究的30种石斑鱼中,鳃棘鲈属(Plectropomus)最先分化,并呈明显单系性;九棘鲈属(Cephalopholis)是一个单系群,并且较石斑鱼属(Epinephelus)原始;侧牙鲈属(Variola)的进化地位介于鳃棘鲈属与九棘鲈属之间;2)宽额鲈(Promicropslanceolatus)可以归入石斑鱼属,而驼背鲈(Cromileptesaltivelis)也与石斑鱼属有很近的亲缘关系,甚至可能是石斑鱼属内的特化类群;3)石斑鱼属内部存在两个平行进化的姐妹分支,分支内部的种类组成与地理分布无关,暗示了石斑鱼属早期的分化模式。The Subfamily Epinephelinae (Teleoste: Serranidae), commonly known as groupers, comprises about 159 species of marine fishes in 15 genera. They are bottom-associated fishes found in tropical and subtropical waters. Most species occur on coral reefs, but some live in estuaries or on rocky reefs. Groupers are of considerable economic value, however, the classification and evolutionary relationships of them have always been hindered by the overwhelming number of species, global distribution, and the lack of morphological specializations traditionally used in ichthyological classification. The confusion and debates have brought much trouble not only to the study of the phylogenetic relationships among groupers, but also to the identification of fry and parent in grouper aquaculture. In this study, to evaluate the phylogenetic relationship of the groupers from the China Seas, the 16S ribosomal DNA of thirty grouper species within six genera of subfamily Epinephelinae were amplified using PCR techniques, 572 bp DNA fragments were obtained and the sequences were analyzed. The genetic information indexes, including Kimura-2 parameter genetic distance and Ts/Tv ratios, were generated using a suite of biology software. Of the 572 aligned base pairs, among which insertion or deletion of base pairs was evident, 373 were constant, 151 were parsimony informative, comprising 26.4% of the total base pairs. The maximum pairwise nucleotide divergence value among all taxa was 0.186 between Epinephelus malabaricus and Plectropomus laevis and the minimum value was 0.007 between E.moara and E.bruneus. The average pairwise nucleotide divergence value among all taxa was 0.079. Transitions were found to be more common than transversions. The average transition/transversion ratio among all pairwise comparisons was 1.7, suggesting that the sequences had not reached saturation. The sequence analysis also showed an obvious adenine bias on the 16S ribosomal DNA fragment and no anti-guanine bias was detected. With Pronotogrammus multifasciatus (No.AF297330) and Johnius belengerii (No.AY336727) as the designated outgroups and additional homologous sequences information of other Cephalopolis and Epinephelus fishes invoked from GenBank, phylogenetic trees were constructed based on the neighbor-joining (NJ),maximum-parsimony (MP) and maximum-likelihood (ML) methods. Conclusions were drawn as follow: 1) all the phylogenic trees suggested that genus Plectropomus, which obviously showed monophyly, is the most primitive group among the subfamily Epinephelinae in this study; genus Cephalopolis is also recovered as a monophyletic group and is more primitive than genus Epinephelus; the evolutionary status of genus Variola is between genus Cephalopolis and genus Plectropomus; 2) Promicrops lanceolatus should be included in genus Epinephelus because it emerged as a cluster within the clade of Epinephelus and had a close relationship with E.coioides and E.malabaricus; Cromileptes altivelis also showed close relationship with genus Epinephelus; perhaps a specialization group within genus Epinephelus; 3) genus Epinephelus was located at the top of the phylogenetic tree indicating that it is the most recently diverged species, which is in accordance with the fact that it is also the most flourishing genus in Epinephelinae. There are two parallel evolutionary sister clades existing (with high bootstrap value support) in genus Epinephelus. However the species-level differences within these clads are not clear due to the lack of high bootstrap values support. The species within this two clades seem to be irrelevant to their biogeographic distributions, which suggested that there were two parallel evolutionary ancestors existing in the early evolution stage of genus Epinephelus and their on-going divergence should be earlier than the existence of geographic floras of genus Epinephelus.国家自然科学基金项目(No.40306023);; 国家自然科学基金项目(No.40576064);; 中国高新技术研究计划(No.2001AA621010)资助~

The identification of Epinephelus malabaricus and E.coioides based on DNA markers

针对在分类上存在混淆的点带石斑鱼和斜带石斑鱼,在形态学基础上,运用mtDNA Cytb基因扩增序列分析、RAPD、AFLP等分子标记技术对两种石斑鱼的遗传关系进行了研究.研究结果表明:(1)点带石斑鱼与斜带石斑鱼的分化已达到种间水平,mtDNA Cytb基因序列相似度为94.4%,AFLP扩增片段的遗传相似率为0.753 9;(2)两种石斑鱼存在杂交现象,RAPD、AFLP的扩增谱图中均获得亲本与子代的特异性条带,父本对杂交子代的遗传影响略大于母本;(3)AFLP标记在点带和斜带石斑鱼及其杂交子代中的遗传符合孟德尔遗传规律,可用于构建两种石斑鱼的遗传连锁图谱.研究结果将对两种石斑鱼种质资源的保护及人工繁育的可续性提供重要参考.Using multi-molecular marker technology and based on morphological criteria,the genetic relationship between E.malabaricus and E.coioides was examined in the hope of resolving the longstanding issue of identifying of these two species.Results showed that:(1) E.coioides and E.malabaricus should be identified as two species,the consistency of mitochondrial DNA cytochrome b gene sequence between E.malabaricus and E.coioides is 94.4%,the genetic similarity by AFLP was 0.753 9;(2)hybridization exists between E.malabaricus and E.coioides,the specific RAPD and AFLP fragments are found to be useful in the identification of these two species,the genetic properties(both with exterior and inheritance) of hybrid is significantly biased to the male parents;and(3)AFLP was a potentially powerful tool in constructing genetic linkage map for these two groupers.国家自然科学基金资助项目(40306023);; 福建省重大专项基金资助项目(2004NZ03

Cloning and Tissue Expression of LECT2 Gene from Epinephelus akaara

基于lECT2的EST序列设计特异引物,采用rACE技术,成功克隆获得赤点石斑鱼lECT2 CdnA全序列,全长601 bP,包括了79 bP的3′uTr区、468 bP的开放阅读框(Orf)以及54 bP的5′uTr区,共编码155个氨基酸.通过与21个物种的氨基酸序列进行同源比对发现,所推导氨基酸序列与大黄鱼的相似性最高,为78%,与其他物种的相似性在40%--73%之间,表明lECT2氨基酸序列具有较高的保守性.基于lECT2基因的氨基酸序列构建的脊椎动物系统进化树与传统物种树基本吻合.同时,通过rT-PCr方法得出,lECT2在健康赤点石斑鱼多种组织中均有表达,肝组织表达量最高,感染哈维氏弧菌(VIbrIO bArVEyI)后lECT2在脾、头肾、鳃等免疫器官中有较高表达量,而在肝脏中表达量上调最为显著,其结果证实了lECT2与赤点石斑鱼免疫应答相关,肝脏可能是赤点石斑鱼lCET2蛋白最主要的表达场所.Based on the sequences from EST library,the full length cDNA of LECT2 was successfully obtained from Epinephelus akaara by RACE technique.The full-length of LECT2 is 601 bp,including 54 bp of 5′UTR,79 bp of 3′UTR,and a 468 bp open reading frame encoding a protein of 155 amino acids.The comparison of LECT2 amino acid sequence in E.akaara with its homologous sequences in other 21 species showed that it shared the highest identity(78%) with LECT2 amino acid sequence in Pseudosciaena crocea,and with the other 20 species,the identity percentages ranged from 40% to 73%.This result indicated that LECT2 is conserved among species.In this study,it was also found that the phylogenetic tree based on LECT2 amino acid sequences is congruent with their evolutionary relationships.Semi-quantitative RT-PCR analyses showed that LECT2 could be detected in various tissues,and the highest expression was found in the liver of E.akaara.Expression of LECT2 in the liver of V.harveyi-challenged fish dramatically increased comparing to that of the healthy fish.Besides the liver,expression in kidney,spleen and gill was slightly higher than those of other examed issues in the V.harveyi-challenged fish.These results revealed that LECT2 might be involved in the process of immunity reactions,and the liver might be the most significant location of LECT2 gene expression.福建省高等学校新世纪优秀人才支持计

Study on Embryonic Development of Two Drumfish Crossbreeding Filial Generagion I

以状黄姑鱼为母本、大黄鱼为父本的人工杂交试验表明 :在温度 17～ 2 0℃、盐度 2 9.9～ 30 .8的条件下 ,杂交卵经 39.9h孵出 ,受精率为 5 8.9% ,孵化率为 1.4% ,仔鱼存活 18d .对照组状黄姑鱼受精率 95 % ,受精卵孵化率 5 6 % .杂交组受精卵的胚胎发育与母本状黄姑鱼的胚胎发育主要差别为 :杂交卵在进入原肠期时 ,多数胚胎发育速度明显变慢 ,囊胚不能正常下包 ,胚胎畸形率较高 .The embryonic development of hybrid combination between Nibea miichthioides ♀ and Pseudosciaena crocea ♂ was studied.The results showed that the oosperm were hatched out in 39.92 h in the state of 17～20 ℃ and 29.9～30.8 of salinity, with the fecundation rate, 59.9% and the hatching rate, 1.4%. The cultivated larvae had been alive for 18 days. As a control group, the fertilization rate of N. miichthioides was 95%, and the hatching rate, 56%. The main difference between the embryonic developments of the crossbreeding and the oosperm of the female parent was likly that the developing speed of the crossbreed was much slower than that of the female parent when they grew to the gastrula, and the blastoderm couldn't develop downward normally, which led to a high proportion of malformations among larvae. The comparison with the male parent and some key factors that affected the intercrossing were also discussed.国家自然科学基金 (39870 5 6 2 );; 国家高技 术发展计划 (86 3 819 0 2 12

Preliminary Study on Induction of Triploidy in Pseudosciaena crocea

以温度休克方法研究了养殖大黄鱼 (Pseudosciaena crocea)三倍体诱导的处理条件 .结果表明 :1)采用冷体克和热休克方法均可诱导出三倍体大黄鱼 ,三倍体诱导率明显受处理温度、处理起始时间以及处理持续时间等因子的影响 ;2 ) 4℃冷休克条件下 ,处理起始时间为受精后 2 min,处理持续时间为 10 min有较高的孵化率和三倍体诱导率 ;3) 37℃热休克条件下 ,处理起始时间为受精后2 .5 min,处理持续时间为 5 min也可获得较高的孵化率和三倍体诱导率Studies on induction of triploidy in the large yellow croaker P. crocea cultured in net cages were respectively carried out from April to September in 2000 in Zhangpu and Ning de. The fertilized eggs were treated with temperature shock at 2~6 ℃ and 37~40 ℃ for 1.5~5 min after fertilization, and the continuation of inducing triploidy was lasted for 5~20 min. The results showed that: when triploidy was induced for 2.5 min after fertilization at 37 ℃ for 5 min, the rate of occurrence of triploid was high; and those for 2 min after fertilization at 4 ℃ for 10 min, the rate of occurrence of triploid was even higher. The relationship between the rate of occurrence of triploid and the survival rate were also discussed.国家高技术研究发展 86 3计划资助项目 (86 3- 819- 0 2 - 12

Analysis on morphological variations among five populations of Marsupenaeus japonicus from coastal areas of China

应用多元分析方法,采用体质量和10个形态性状为指标对中国沿海陵水、北海、惠来、诏安、厦门5个地理群体的日本囊对虾(MArSuPEnAEuS JAPOnICuS)进行了比较分析,结果表明,日本囊对虾雌雄个体间形态差异显著(P0.05);雌性个体的体长、头胸甲宽和雄性个体的体长、第六腹节宽分别与体质量的相关系数达到了极显著水平(P<0.01);雌性日本囊对虾的体长(X2)、头胸甲宽(X4)对体质量(y)的最优多元回归方程为:y=-151.737+9.235X2+23.194X4,雄性日本囊对虾体长(X2)、第六腹节宽(X9)对体质量(y)的最优多元回归方程为:y=85.280+5.222X2+43.656X9。以上结果为日本囊对虾的选择育种提供了理想的测度指标。Body weight and ten morphological traits in five geographic groups of Marsupenaeus japonicus in coastal areas of China were compared using multivariate analysis method.Results show that there were significant variations between male and female M.japonicus while no significant variations between different geographic populations.For the same gender,populations from Hainan and Beihai were closely linked and those from Xiamen and Zhaoan were closely linked.Significant correlation coefficients(P<0.01)were found between body weight(Y)and morphological traits of body length(X 2 )and carapace width(X 4 )for females,and between body weight(Y)and body length(X 2 ) and the sixth abdominal segment width(X 9 )for males,which are ideal indicators of measurement for M.japonicus aquaculture.The multiple regression equations were establised as Y=-151.737+9.235X 2 +23.194X 4 for females and Y=85.280+5.222X 2 +43.656X 9 for males.农业部公益性行业(农业)科研专项资助项目(200803012);福建省科技厅重大项目(2009N3011