تشخیص، بررسی فیلوژنی و کلونینگ ژن G گلیکوپروتئین ویروس سپتی سمی هموراژیک (VHSV) جدا شده از ماهی قزل آلای رنگین کمان (Onchorhynchus mykiss)

نوع مقاله : بیماری ها

نویسندگان

1 گروه بهداشت و بیماری های آبزیان، دانشکده دامپزشکی، دانشگاه شیراز، شیراز، ایران

2 گروه پاتوبیولوژی، دانشکده دامپزشکی، دانشگاه شیراز، شیراز، ایران

3 گروه تشخیص بیماری های آبزیان، مرکز ملی تشخیص، آزمایشگاه‌های مرجع و مطالعات کاربردی، سازمان دامپزشکی کشور، تهران، ایران

چکیده

بیماری سپتی ­سمی خونریزی ­دهنده از مهم­ترین بیماری ­های ویروس آزادماهیان می ­باشد. ژنوم این ویروس شامل شش ژن می­ باشد که مهم ­ترین آن­ ژن G (گلیکوپروتئین) است و در ایمنی ­زایی علیه این ویروس نقش دارد. هدف از این مطالعه شناسایی ویروس سپتی ­سمی خونریزی ­دهنده قزل­ آلای رنگین­ کمان، مطالعه فیلوژنی و کلونینگ ژن G ویروس در وکتور pTZ57r/t بود. پس از تشخیص ویروس سپتی ­سمی خونریزی­ دهنده به ­روش RT-PCR و کلونیگ ژن G داخل پلاسمید pTZ57r/t، پلاسمید نوترکیب سنتزشده به باکتری پذیرا (E.coli) منتقل گردید. از کلونی ­های سفید دربردارنده وکتور نوترکیب جهت استخراج پلاسمید استفاده شد. اندازه ژن G ویروس سپتی­ سمی هموراژیک bp 1523 می ­باشد که در این پژوهش جهت تایید کلونینگ از روش کلونی PCRو تعیین توالی اسفاده شد. بررسی­ های حاصل از این مطالعه نشان داد که ژن G با موفقیت در وکتور  pTZ57r/t کلون گردیده است و می­ توان ازآن جهت بیان گلیکوپروتئین ویروس در وکتورهای بیانی به ­منظور تولید پروتئین استفاده کرد. نتایج حاصل شده از تعیین توالی ژن G نیز نشان داد که ویروس شناسایی شده در این مطالعه متعلق به ژنوتیپ Ia-2  بوده است. مطالعه حاضر نخستین تحقیقی است که در کشور به ­منظور کلونینگ ژن Gویروس سپتی­ سمی خونریزی ­دهنده در وکتورهای پلاسمیدی انجام شده است.

کلیدواژه‌ها

عنوان مقاله [English]

Diagnosis, Phylogenetic analysis and cloning of G glycoprotein gene of viral hemorrhagic septicemia virus (VHSV) detected in Rainbow trout (Onchorhynchus mykiss)

نویسندگان [English]

  • Amir Ali Heidari 1
  • Mostafa Akhlaghi 1
  • Ali Mohammadi 2
  • laleh Moazemi goudarzi 3
  • Azadeh Yektaseresht 2
1 Department of Aquatic Health and Diseases, Faculty of Veterinary, Shiraz University, Shiraz, Iran
2 Department of Pathobiology, Faculty of Veterinary, Shiraz University, Shiraz, Iran
3 Aquatic Diseases Diagnosis Group, National Center for Diagnosis, Reference Laboratories and Applied Studies, Veterinary Organization, Tehran, Iran
چکیده [English]

Viral hemorrhagic septicemia is one of the most important virus disease in salmonid fishes. The genome of the virus consists of six genes, the most important of which is the G glycoprotein gene, which plays a role in immunization against the virus. The aim of this study was detection of viral hemorrhagic septicemia virus in Rainbow trout, phylogenic study and cloning of the G gene of virus in pTZ57r/t vector. Methods: After detecting of viral hemorrhagic septicemia virus by RT-PC, cloning of G gene in pTZ57r/t plasmid, the recombinant plasmid was transformed to the competent bacterium (E. coli). White clonies containing recombinant vector were used to extract the plasmid. The size of the G gene of viral hemorrhagic septicemia virus was 1523 bp. In this study, colony PCR method and sequencing were used to confirm the cloning. Results: The results of this study showed that G gene has been cloned successfully in pTZ57r/t vector and can be used to express the glycoprotein of the virus in expression vectors for protein production. The results of sequencing of G gene also showed that the virus identified in this study belonged to genotype Ia-2.

کلیدواژه‌ها [English]

  • VHSV
  • G glycoprotein
  • Rainbow trout
  • Cloning

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فصلنامه محیط زیست جانوری
دوره 12، شماره 2
تیر 1399
صفحه 145-150

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