凡纳滨对虾CTSL基因PCR-SSCP多态性与生长 性状的关联分析及其mRNA的差异表达
PCR-SSCP Polymorphism of CTSL gene and its correlation with growth traits of Litopenaeus vannamei and the different mRNA expressions of CTSL
-
摘要: 利用PCR-SSCP技术对379尾凡纳滨对虾组织蛋白酶Cathepsin L(CTSL)基因多态性进行检测,采用最小二乘法对多态性与生长性状的相关性进行分析;利用Real-time quantitative PCR检测CTSL基因mRNA的差异表达情况。结果表明:引物C6 (F/R)扩增片段具有多态性,其扩增产物具有CC、CT和TT 3种基因型,TT基因型与CC基因型相比在第6外显子92 bp处发生了C→T的突变,为沉默突变。最小二乘法分析结果表明,在体重、体长、第一腹节长和尾节长这4个指标上CC和CT基因型显著高于突变型TT(p<0.05),而在头胸甲长和头胸甲宽2个指标上,CC基因型显著高于CT和TT基因型(p<0.05)。CTSL基因mRNA在凡纳滨对虾各组织、不同性别个体肌肉组织、极端体重个体肌肉组织中相对表达量大小顺序分别为:肝胰脏>胃>眼柄>心脏>消化腺;雌虾>雄虾;极大个体>极小个体。结论:CTSL基因多态性对凡纳滨对虾生长性状有显著影响,可以作为影响该虾生长性状的候选基因,为该虾的选育提供分子遗传标记;CTSL基因mRNA的分布情况表明此基因可能正向调控对虾生长。Abstract: The polymorphism of CTSL gene in 379 Litopenaeus vannamei was detected using PCR-SSCP technique and its correlations with growth traits of L.vannamei was analyzed by least square method in this study. In addition, differential mRNA expressions of CTSL were carried out by real-time quantitative PCR. The results showed that the product amplified by primer C6 (F/R) displayed polymorphisms containing three genotypes: CC, CT and TT. Compared with the CC genotype, a silence mutation (C→T) occurred in TT genotype in exon6(92 bp)of the CTSL gene. For body mass, body length, the first abdominal segment length, and the last abdominal segment length, shrimps of CC and CT genotypes were significantly higher than those of the TT genotype(p<0.05), while for carapace length and carapace width, shrimps of CC genotype were significantly higher than those of CT and TT(p<0.05). The expressions of CTSL mRNA in various tissues, muscle tissues of different sex, different body weight individuals were: hepatopancreas>stomach>eyestalk>heart>digestive gland, female>male and heavy>light, respectively. The results obtained in this study suggest CTSL gene may be one of the candidates which significantly affect the growth of Litopenaeus vannamei. It can be used as the molecular genetic marker applying for Litopenaeus vannamei breeding. The distribution of the CTSL mRNA may be related to its specific functions in different tissues and its positive roles in the regulation of the shrimp muscle growth.
-
Key words:
- Litopenaeus vannamei /
- Cathepsin L gene /
- PCR-SSCP /
- correlation analysis /
- differential expression
-
周鑫. 南美白对虾的苗种繁育与淡水养殖技术[J]. 科学养鱼, 2003:1-15. 吴立峰, 张吕平, 沈琪, 等. 凡纳滨对虾不同家系的形态性状对体重的影响[J]. 海洋湖沼通报, 2010 (2) :37-48. 张存善, 李色东, 王亚平, 等. 2个凡纳滨对虾群体仔虾的生长比较[J]. 水产科学, 2010, 29 (06) :325-328. Senapin S, Thaowbut Y, Gangnonngiw W, et al. Impact of yellow head virus outbreaks in the white leg shrimp, Penaeus vannamei (Boone) , in Thailand[J]. Journal of Fish Diseases, 2010, 33 (5) :421-430. Lic C, Yeh S T, Chen J C. Innate immunity of the white shrimp Litopenaeus vannamei weakened by the combination of a Vibrio alginolyticus injection and low salinity stress[J]. Fish & Shelfish Immunology, 2010, 28 (1) :121-127. Schuler D J, Boardman G D, Kuhn D D, et al. Acute toxicity of ammonia and nitrite to Pacific white shrimp, Litopenaeus vannamei, at low salinities[J]. Journal of the World Aquaculture Society, 2010, 41 (3) :438-446. Page R D M. Treeview: An application to display phylogenetic trees on personal computers[J]. Comp Appt Bioscl, 1996, 12:357-358. Schneider S, Kueffer J M, Roessli D, et al. A software for population genetic data analysis[M]. Switzerland:Genetics and Biometry Laboratory, University of Geneva, 1997. Rice R W. Analyzing table of statistical tests[J]. Evolution, 1989, 43:223-225. 李峰, 刘楚吾, 林继辉. 两个凡纳滨对虾亲本群体的RAPD研究[J]. 湛江海洋大学学报, 2003, 23 (4) :1-5. Zhang L, Yang C, Zhang Y, et al. A genetic linkage map of Pacific white shrimp (Litopenaeus vannamei) :Sex-linked microsatellite markers and high recombination rates[J]. Genetica, 2006 Oct 17: Epub ahead of print. Zeng D, Chen X, Li Y, et al. Analysis of Hsp70 in Litopenaeus vannamei and detection of SNPs[J]. Crustacean Biology, 2008, 28 (4) :727-730. 辛静静, 刘小林, 李喜莲, 等.凡纳滨对虾α-淀粉酶基因PCR-RFLP多态性与生长性状的相关分析[J]. 海洋学报, 2011, 33 (3) :124-130. Du Z Q, Ciobanu D C, Onteru S K, et al. A gene-based SNP linkage map for Pacific white shrimp, Litopenaeus vannamei[J]. Animal genetics, 2010, 41 (3) :286-294. Glenn K L, Grapes L, Suwanasopee T, et al. SNP analysis of AMY2 and CTSL genes in Litopenaeus vannamei and Penaeus monodon shrimp[J].Anim Genet, 2005 Jun, 36 (3) : 235-236. Aljanabis M, Martinez I. Universal and rapid salt extraction of high quality genomic DNA for PCR-based techniques[J]. Nucleic Acids Reserch, 1997, 25 (22) : 4692-4693. Kenneth J L, Thomas D S. Analysis of Relative Gene Expression Data Using Realtime Quantitative PCR and the 2-△△CT Method[J]. Methods, 2001, 25: 402-408. Xie P, Liu J K. Phytoplankon, especially diatoms, in the gut contents and feces of two plantivorous cyprinds silver carp and bighead carp[J]. China Oceanol Limnol, 1994, 12: 308-315.
点击查看大图
计量
- 文章访问数: 1634
- HTML全文浏览量: 17
- PDF下载量: 924
- 被引次数: 0