Protection of Mice Against Challenge with Foot-and-Mouth Disease Virus by Immunization with vp1 Gene Expressed Transgenic N·Benthamiana
Abstract: A binary plasmid was constructed by triparental mating, which contained the vp1 gene of foot-and-mouth disease virus strain Akesu/58 serotype O. The Nicotiana Benthamiana (NC) leaves were transformed by Agrobacterium tumefaciens-mediated method with the binary expression plasmid. After selected by kanamycin, forty-nine resistant lines were obtained. Forty lines were positive through PCR with total DNA and twenty-one lines were positive through RT-PCR with total RNA from fresh leaves tissue. The leave extracts of seven positive lines analyzed with ELISA and western-blot were respectively emulsified in Freund′s adjuvant and the Balb/C mice were immunized intraperitoneally at days 0,15,30 and 45. After the last inoculation, the sera antibody were analyzed at 9 days and the mice were experimentally challenged intraperitoneally with 10~4 suckling mouse 50% lethal doses (SM_(50)LD) of FMDV at 12 days. Protection was determined by the absence of viremia at 24h post-inoculation. ˎ̥,Verdana,Arial; mso-font-kerning: 1.0pt; mso-bidi-font-family: 'Times New Roman'; mso-fareast-font-family: 宋体; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA">Viremia was tested by intradermal in ˎ̥,Verdana,Arial; mso-font-kerning: 1.0pt; mso-bidi-font-family: 'Times New Roman'; mso-fareast-font-family: 宋体; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA">intradermal inoculation of perip heral blood into suckling mice. These results showed that the const ructed binary expression plasmid was functionary ; vp1 gene was transformed into NC plants and expresed affirmatively ; the structure protein VP1 expressed in two transgenic plants lines may possibly be used as a source of antigen , the inoculated Balb/C mice developed a specific antibody response ; the protection against FMDV challenge in two groups was 100 % and 63 % respectively.