| ZHU Yanbing,YANG Feng. 2007. Construction of white spot syndrome virus (WSSV) whole genome phage display library. Acta Oceanologica Sinica, (2):75－83 |
| Construction of white spot syndrome virus (WSSV) whole genome phage display library |
| Construction of white spot syndrome virus (WSSV) whole genome phage display library |
| Received:March 09, 2006 Revised:July 20, 2006 |
| DOI： |
| Key words:white spot syndrome virus genome phage display library dot blot |
| 中文关键词: white spot syndrome virus genome phage display library dot blot |
| 基金项目: |
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| Abstract: |
| A rebuilt vector pCANTAB 5 EE was obtained by inserting a 34 bp double-stranded oligonucleotide which contained a EcoRV recognition sequence into pCANTAB 5 E.White spot syndrome virus (WSSV) genome DNA was fragmented by sonication to isolate fragments mainly in the range of 0.8~2.0 kb, then the fragments were blunt-ended with T4 DNA polymerase and cloned into the EcoRV site of pCANTAB 5 EE.The primary recombinant clone of the library was 3.0×105.Colony PCR of random selected recombinants showed that the size of the inserts was 0.12~1.77 kb.After the whole library recombinant phages infected Escherichia coli HB2151 cells, the extracellular and periplasmic extracts were dropped on PVDF membranes to perform dot blot, using polyclonal mouse anti-VP24 serum, anti-WSV026 serum, anti-WSV063 serum, anti-WSV069 serum, anti-WSV112 serum, anti-WSV238 serum, anti-WSV303 serum and anti-VP26 serum as the primary antibody, respectively.The results showed that the display library could express the viral proteins. |
| 中文摘要: |
| A rebuilt vector pCANTAB 5 EE was obtained by inserting a 34 bp double-stranded oligonucleotide which contained a EcoRV recognition sequence into pCANTAB 5 E.White spot syndrome virus (WSSV) genome DNA was fragmented by sonication to isolate fragments mainly in the range of 0.8~2.0 kb, then the fragments were blunt-ended with T4 DNA polymerase and cloned into the EcoRV site of pCANTAB 5 EE.The primary recombinant clone of the library was 3.0×105.Colony PCR of random selected recombinants showed that the size of the inserts was 0.12~1.77 kb.After the whole library recombinant phages infected Escherichia coli HB2151 cells, the extracellular and periplasmic extracts were dropped on PVDF membranes to perform dot blot, using polyclonal mouse anti-VP24 serum, anti-WSV026 serum, anti-WSV063 serum, anti-WSV069 serum, anti-WSV112 serum, anti-WSV238 serum, anti-WSV303 serum and anti-VP26 serum as the primary antibody, respectively.The results showed that the display library could express the viral proteins. |
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