AsiaⅠ型口蹄疫基因治疗与基因免疫双功能疫苗载体的构建
摘要
目前,对于口蹄疫的防制主要采用屠杀、隔离和免疫接种,其中疾病流行前的免疫接种是特异性保护家畜免遭感染的有效途径,但现有疫苗接种动物后都会存在免疫空白期,即诱导免疫动物产生体液和细胞免疫反应,等待有效抗体产生的一段时间。动物如果在这段时间内遭到病毒的入侵往往会引起该病的流行。为探索解决免疫动物在疫苗接种的免疫空白期内发病这一难题,本研究以Asia I江苏株为模板,引入RNA干扰机制,进行了双功能疫苗的探索研究,并取得下述进展:
1.对pBudCE4.1载体进行改造,将其CMV启动子替换为U6启动子,成功改造出适合于双功能疫苗研究的双启动子载体pBudCE4.1-U6。
2.利用绿色荧光蛋白的表达与沉默表达对载体pBudCE4.1-U6的双启动子分别进行功能效率鉴定,结果表明对载体的改造是成功的,载体pBudCE4.1-U6的两个启动子均能启动目的基因的表达。
3.针对Asia I江苏株的3D基因设计3条小RNA干扰片段,并将之与免疫组合基因P12A+3C克隆入双启动子载体pBudCE4.1-U6,成功构建出了3个双功能疫苗质粒。
4.双功能疫苗质粒转染靶细胞BHK-21,有延迟细胞病变的作用,说明双功能载体能够转录出siRNA,对病毒的转录复制有一定的抑制效果。
5.间接免疫荧光检测结果表明双功能载体能够同时启动口蹄疫免疫基因在BHK-21细胞中的表达。
At present, slaughter, isolation and vaccination are major methods to control FMD, and vaccination before epidemic of FMD is the specific efficient method to protect animals against virus entry. But a challenge to the development of vaccines against microbiology is the exist of immunization black and the latent period, FMD would outbreak when animals challenged with virus during the immunization black as well as animals inoculate the vaccine during the latent period. In order to solve this problem, we introduced RNAi into the bi-functional vaccine research, using Asia I/JS strain as the model to confirm the feasibility of the conception.
1. Replaced the CMV promoter of vector pBudCE4.1 by U6 promoter, developed a dual-promoter vector pBudCE4.1-U6.
2. Tested the functions of the promoters of vector pBudCE4.1-U6 by expressing EGFP and Interfering EGFP expression. Results demonstrated that both CMV promoter and U6 promoter could play its role.
3. We successfully constructed three bi-functional plasmids expressing shRNAs targeting 3D gene of Asia I/JS strain FMDV and expressing immunization protein P12A+3C simultaneously.
4. The antiviral potential induced by bi-functional plasmid was evident in the face of challenge of with a homologous FMDV and the inhibition was inconspicuousness.
5. Detection of indirect immunofluorescence showed that immunization protein was expressed in BHK-21 cells.
1.对pBudCE4.1载体进行改造,将其CMV启动子替换为U6启动子,成功改造出适合于双功能疫苗研究的双启动子载体pBudCE4.1-U6。
2.利用绿色荧光蛋白的表达与沉默表达对载体pBudCE4.1-U6的双启动子分别进行功能效率鉴定,结果表明对载体的改造是成功的,载体pBudCE4.1-U6的两个启动子均能启动目的基因的表达。
3.针对Asia I江苏株的3D基因设计3条小RNA干扰片段,并将之与免疫组合基因P12A+3C克隆入双启动子载体pBudCE4.1-U6,成功构建出了3个双功能疫苗质粒。
4.双功能疫苗质粒转染靶细胞BHK-21,有延迟细胞病变的作用,说明双功能载体能够转录出siRNA,对病毒的转录复制有一定的抑制效果。
5.间接免疫荧光检测结果表明双功能载体能够同时启动口蹄疫免疫基因在BHK-21细胞中的表达。
At present, slaughter, isolation and vaccination are major methods to control FMD, and vaccination before epidemic of FMD is the specific efficient method to protect animals against virus entry. But a challenge to the development of vaccines against microbiology is the exist of immunization black and the latent period, FMD would outbreak when animals challenged with virus during the immunization black as well as animals inoculate the vaccine during the latent period. In order to solve this problem, we introduced RNAi into the bi-functional vaccine research, using Asia I/JS strain as the model to confirm the feasibility of the conception.
1. Replaced the CMV promoter of vector pBudCE4.1 by U6 promoter, developed a dual-promoter vector pBudCE4.1-U6.
2. Tested the functions of the promoters of vector pBudCE4.1-U6 by expressing EGFP and Interfering EGFP expression. Results demonstrated that both CMV promoter and U6 promoter could play its role.
3. We successfully constructed three bi-functional plasmids expressing shRNAs targeting 3D gene of Asia I/JS strain FMDV and expressing immunization protein P12A+3C simultaneously.
4. The antiviral potential induced by bi-functional plasmid was evident in the face of challenge of with a homologous FMDV and the inhibition was inconspicuousness.
5. Detection of indirect immunofluorescence showed that immunization protein was expressed in BHK-21 cells.
引文
[1]陈煜,谢小芳. RNAi的作用机制及抗病毒研究进展.世界华人消化杂志, 2006, 14(21): 2123-2129.
[2] Yu JY, DeRuiter SL, Turner DL. RNA interference by expression of short-interfering RNAs and hairpin RNAs in mammalian cells[J]. Proc Natl Acad Sci, 2002, 99(9): 6047-6052.
[3] Novina C.D, and Sharp P.A. The RNAi revolution. Nature 2004; 430: 161-164.
[4] Jacque JM, Triques K , Stevenson M. Modulation of HIV21 replication by RNA interference[J] . Nature , 2002, 418: 435-438.
[5] Ge Q , Filip L , Bai A ,et al. Inhibition of influenza virus production in virus infected mice by RNA interference[J]. Proc Natl Acad Sci USA, 2004, 101 (23): 8676-8681.
[6] Chen W, Yan W, Du Q,et al. RNA interference targeting VPI inhibits foot and mouth disease virus replication in BHK21 cells and suckling mice[J]. J Virol, 2004, 78 (13): 6900-6907.
[7] Hu WY, Myers CP, Kilzer JM,et al. Inhibition of retroviral pathogenesis by RNA interference[J]. Curr Biol, 2002, 12: 1301-1311.
[8] Jiang M, Milner J. Selective silencing of viral gene expression in HPV2 positive human cervical carcinoma cells treated with siRNAs,a primer of RNA interference[J]. Oncogene, 2002, 21: 6041-6048.
[9] Sui G, Soohoo C, Affar E B, et al. A DNA vector based RNAi technology to suppress gene expression in mammalian cells[J]. Genetics, 2002, 99 (8): 5515-5520.
[10]韩晓荣,柳纪省,杨彬等.亚洲1型口蹄疫病毒反义核酸双向表达载体的构建[J].动物医学进展,2008, 29(3): 5-9.
[11]谢书阳,张敬之,黄淑帧等.应用RNAi抑制细胞中绿色荧光蛋白的表达[J].中华医学遗传学杂志, 2005, 22(4): 431-434.
[12] Coumoul X, Deng CX. RNAi in mice: a promising approach to decipher gene functions in vivo[J]. Biochimie,2006,88(6): 637-643.
[13] JN Leonard, DV Schaffer Antiviral RNAi therapy: emerging approaches for hitting a moving target[J]. Gene Therapy, 2006, 13: 532-540.
[14] Seon-Young Kima, Jae-Ho Leea, Hyun-Seock Shina,et al. The human elongation factor 1 alpha (EF-1α) first intron highly enhances expression of foreign genes from the murine cytomegalovirus promoter[J]. Journal of Biotechnology, 2002, 93(2): 183-187.
[15] Vivek Shukla, Xavier Coumoul, Chu-Xia Deng. RNAi-based conditional gene knockdown in
[1]陈煜,谢小芳. RNAi的作用机制及抗病毒研究进展.世界华人消化杂志, 2006, 14(21): 2123-2129.
[2] Yu JY, DeRuiter SL, Turner DL. RNA interference by expression of short-interfering RNAs and hairpin RNAs in mammalian cells[J]. Proc Natl Acad Sci, 2002, 99(9): 6047-6052.
[3] Novina C.D, and Sharp P.A. The RNAi revolution. Nature 2004; 430: 161-164.
[4] Jacque JM, Triques K , Stevenson M. Modulation of HIV21 replication by RNA interference[J] . Nature , 2002, 418: 435-438.
[5] Ge Q , Filip L , Bai A ,et al. Inhibition of influenza virus production in virus infected mice by RNA interference[J]. Proc Natl Acad Sci USA, 2004, 101 (23): 8676-8681.
[6] Chen W, Yan W, Du Q,et al. RNA interference targeting VPI inhibits foot and mouth disease virus replication in BHK21 cells and suckling mice[J]. J Virol, 2004, 78 (13): 6900-6907.
[7] Hu WY, Myers CP, Kilzer JM,et al. Inhibition of retroviral pathogenesis by RNA interference[J]. Curr Biol, 2002, 12: 1301-1311.
[8] Jiang M, Milner J. Selective silencing of viral gene expression in HPV2 positive human cervical carcinoma cells treated with siRNAs,a primer of RNA interference[J]. Oncogene, 2002, 21: 6041-6048.
[9] Sui G, Soohoo C, Affar E B, et al. A DNA vector based RNAi technology to suppress gene expression in mammalian cells[J]. Genetics, 2002, 99 (8): 5515-5520.
[10]韩晓荣,柳纪省,杨彬等.亚洲1型口蹄疫病毒反义核酸双向表达载体的构建[J].动物医学进展,2008, 29(3): 5-9.
[11]谢书阳,张敬之,黄淑帧等.应用RNAi抑制细胞中绿色荧光蛋白的表达[J].中华医学遗传学杂志, 2005, 22(4): 431-434.
[12] Coumoul X, Deng CX. RNAi in mice: a promising approach to decipher gene functions in vivo[J]. Biochimie,2006,88(6): 637-643.
[13] JN Leonard, DV Schaffer Antiviral RNAi therapy: emerging approaches for hitting a moving target[J]. Gene Therapy, 2006, 13: 532-540.
[14] Seon-Young Kima, Jae-Ho Leea, Hyun-Seock Shina,et al. The human elongation factor 1 alpha (EF-1α) first intron highly enhances expression of foreign genes from the murine cytomegalovirus promoter[J]. Journal of Biotechnology, 2002, 93(2): 183-187.
[15] Vivek Shukla, Xavier Coumoul, Chu-Xia Deng. RNAi-based conditional gene knockdown in
[34] Davenport D, Nichol JAC: Luminescence in Hydromedusae. Proceedings of the Royal Society, Series B 1955, 144:399-411.
[35] Morin JG, Hastings JW: Energy transfer in a bioluminescent system. Journal of Cellular Physiology 1971, 77:313-318.
[36] Anderson JM, Cormier MJ: Lumisomes, the cellular site of bioluminescence in coelenterates. The Journal of Biological Chemistry 1973, 248:2937-2943.
[37] Morise H, Shimomura O, Johnson FH, Winant J: Intermolecular energy transfer in the bioluminecent system of Aequorea. Biochemistry 1974, 13:2656-2662.
[38] Prasher DC, Eckenrode VK, Ward WW, Prendergast FG, Cormier MJ: Primary structure of the Aequorea victoria green fluorescent protein. Gene 1992, 111:229-233.
[39] Bokman SH, Ward WW: Renaturation of Aequorea green fluorescent protein. Biochemical and Biophysical Research Communications 1981, 101:1372-1380.
[40] Cody CW, Prasher DC, Westler WM, Prendergast FG, Ward WW: Chemical-structure of the hexapeptide chromophore of the Aequorea green fluorescent protein. Biochemistry 1993, 32:1212-1218.
[41] Heim R, Prasher DC, Tsien RY: Wavelength mutations and post-translational autoxidation of green fluorescent protein. Proceedings Of the National Academy Of Sciences Of the United States Of America 1994, 91:12501-12504.
[42] Chalfie M, Tu Y, Euskirchen G, Ward WW, Prasher DC: Green fluorescent protein as a marker for gene expression. Science 1994, 263:802-805.
[43] Wang SX, Hazelrigg T: Implications for bcd mRNA localization from spatial distribution of exu protein in Drosophila oogenesis. Nature 1994, 369:400-403.
[44] Baulcombe DC, Chapman S, Cruz SS: Jellyfish green fluorescent protein as a reporter for virus infections. Plant Journal 1995, 7:1045-1053.
[45] Heinlein M, Epel BL, Padgett HS, Beachy RN: Interaction of tobamovirus movement proteins with the plant cytoskeleton. Science 1995, 270:1983-1985.
[46] Haseloff J, Amos B: GFP in plants. Trends in Genetics 1995, 11:328-329.
[47] Hu W, Cheng CL: Expression of Aequorea green fluorescent protein in plant cells. FEBS Letters 1995, 369:331-334.
[48] Sheen J, Hwang SB, Niwa Y, Kobayashi H, Galbraith DW: Green fluorescent protein as a new vital marker in plant cells. Plant Journal 1995, 8:777-78.
[49] Siemering, K.R. Golbik, R., Sever, R. & Haseloff, J. Mutations that supress the thermosensitivity of green fluorescent protein. Current Biology 6:1653-1663, 1996
[50] Haseloff. J. Siemering, K.R., Prasher, D. & Hodge, S. Removal of a cryptic intron and subcellular localisation of green fluorescent protein are required to mark transgenic Arabidopsis plants brightly. Proc. Natl. Acad. Sci. USA, in press.
[51]卢曾军,刘在新.口蹄疫病毒研究进展[J].中国兽医科技, 2003, 33(2):69- 74.
[52]蔡宝祥.家畜传染病学.北京:中国农业出版社,1999. 104 -111.
[53] Carroll A R, Rowlands D J, Carke B E. The complete nucleotide sequence of the RNA coding for the primary translation product of foot-and-mouth disease virus[J].Nucleic Acids Res ,1984 ,12 (5) :2461 -2472.
[54] Forss S, Strebel K, Beck E, et al, Nucleotide sequence and genome organization of Foot-and-mouth disease virus [J]. Nucleic Acids Res, 1984, 12: 6587 - 66011
[55] Pilipenko E V, Blinov V M, Dmitrieva T M. Conservation of the secondary structure elements of 5’-untranslated region cardio and aphthovirus RNAs [J]. Nucleic Acids Research, 1989, 17(14):5701-5711.
[56] Stassinopoulos I A, Belsham G J. A novel protein RNA binding assay: functional interactions of the foot-and-mouth disease virus internal ribosome entry site with cellular proteins [J]. RNA, 2001, 7(1):114-122.
[57] Ohlmann T,Jackson R J.The properties of chimeric picornavirus IRESes show that internal translation initiation sites is influenced by the identity of the IRES and not just the context of the AUG codon[J]. RNA, 1999, 5(6):764-778.
[58] Lopez de Quinto S, Lafuente E, Martinez Salas E. IRES interaction with translation initiation factors: functional characterization of novel RNA contacts with eIF3, eIF4B, and eIF4GII[J]. RNA, 2001, 7(9):1213-1226.
[59] Niepmann M. Effects of potassium and chloride on ribosome association with the RNA of foot-and-mouth disease virus [J]. Virus Res, 2003, 93(1):71-78.
[60]张显升,赵启祖,刘在新,等.口蹄疫病毒基因组内部核糖体进入位点一级和二级结构分析[J].病毒学报, 2002, 18(2):142-148.
[61] Sagedahl A, Giraudo A T, Mello P, et al. Biochemical characterization of an aphthovirus typeC strain Resende attenuated for cattleby serial passages in chicken embryos [J]. Virology, 1987, 157: 366- 374
[62] Gutierrez A, Martinez-Selas E, Pintado B, et al. Specific inhibition of aphthovirus infection by RNAs transcribed from both the 5’and 3’noncoding regions[J].J Viral, 1994, 68 (11): 7436– 7438.
[63] Mateu M G. Antibody recognition of picornaviruses and escap from neutralization: a structural view [J]. Virus Rec, 1995, 38: 1-24.
[64] Wu Q, Moraes M P, Grubman M J. Recombinant adenovirus co-expressing capsid proteins of two serotypes of foot-and-mouth disease virus (FMDV): in vitro characterization and induction of neutralizing antibodies against FMDV in swine [J]. Virus Res, 2003, 93 (2):211-219.
[65] Tosh C, Hemadri D, Sanyal A, et al. Genetic and antigenic analysis of two recently circulating genotypes of type a foot-and-mouth disease virus in India: evidence for positive selection in the capsid coding genes [J]. Arch Virol, 2003, 148(5):853-869.
[66] Balamurugan V, Renji R, Saha S N,et al. Protective immune response of the capsid precursor polypeptide (P1) of foot and mouth disease virus type O produced in Pichia pastoris[J]. Virus Res, 2003, 92(2):141-149.
[67]赵启祖,谢庆阁.口蹄疫病毒抗原位点的研究及应用[A].畜禽重大疫病免疫防制研究进展[C].谢庆阁,翟中和.北京:中国农业出版社,1996,14-23.
[68]徐为燕.兽医病毒学[M].北京:农业出版社,1993:56-59.
[69] Kokuho T, Watanabe S, Yokamizo Y, et al. Production of biologically active,heterodimeric porcine interleukin using a monocistromic baculoviral expression system[J]. Vet Immunol Immunopathol, 1999, 72(3, 4): 289-302.
[70] Gromeier M, Wimmer E, Gorbalenga A E. Genetics, pathogensis and evolution of piconaviru- ses [A]. Domingo E, Webster R, Holland J, et al. Origin and evolution of viruses[C]. Academic Press,1999:287- 343.
[71]江鹏斐,谢庆阁.口蹄疫研究进展[A].谢庆阁,翟中和.兽医生物制品基础研究[C].北京:中国农业科技出版社,1996: 1- 10.
[72] Beard C W, Mason P W. Genetic determinants of altered virulence of Taiwanese foot-and-mouth disease virus [J]. Virol, 2000, 71:987-991.
[73] Knowles N, Davies P R, Henry T, et al. Emergence in Asia of foot-and-mouth disease viruses with altered host range: characterization of alteration in the 3A protein [J]. Virol, 2001, 75(3): 1551-1556.
[74] Wolff J.A., Malone R. W., Williams P, et al. Direct gene transfer into mouse muscle invivo. Science, 1990, 247:1465-1468.
[75]张显升.口蹄疫病毒基因组RNA结构与功能研究进展.病毒学报, 2001, l7(4):375-380.
[76] Grubman MJ, Baxt B.Foot-and-Mouth Disease [J].Clin.Microbiol.Rev. 2004,17:465-493.
[77] DomingoE,EscarmisC,Baranowski E,et al.Evolution of foot-and-mouth disease virus[J].Virus Research, 2003, 91(1):47-63.
[78]农业部畜牧兽医局.一、二、三类动物疫病释义[M].北京:中国农业出版社,2004,1-5.
[79] Guo H, Liu X, Liu Z, et al. Recent outbreaks of foot-and-mouth disease type Asia I in China [J]. J Vet Med B Infect Dis Vet Public Health, 2006, 53(S1): 29-33.
[80]田赠义,尹德华.口蹄疫防疫技术[M].兰州:甘肃民族出版社,2001: 122.
[81]张怀宇.口蹄疫灭活疫苗质量控制和检测技术的研究进展.兽药导刊, 2007: 26-29.
[82]徐全武,申涛.口蹄疫疫苗研究进展[J].新疆畜牧业.2003(3):47-48.
[83]农业部畜牧兽医司.家畜口蹄疫及其防治[M].北京:中国科技出版社,1994:181-184.
[84]薛青红,刘淋涛.口蹄疫病毒3A基因在大肠杆菌中的高效表达[J].微生物学报,2002,5(4): 540 -542.
[85] Bin Yang, Xi Lan, Xueri Li,et al. A novel bi-functional DNA vaccine expressing VP1 protein and producing antisense RNA targeted to 5′UTR of foot-and-mouth disease virus can induce both rapid inhibitory effect and specific immune response in mice[J]. Vaccine, 2008, 26(43), 5477-5483.
[2] Yu JY, DeRuiter SL, Turner DL. RNA interference by expression of short-interfering RNAs and hairpin RNAs in mammalian cells[J]. Proc Natl Acad Sci, 2002, 99(9): 6047-6052.
[3] Novina C.D, and Sharp P.A. The RNAi revolution. Nature 2004; 430: 161-164.
[4] Jacque JM, Triques K , Stevenson M. Modulation of HIV21 replication by RNA interference[J] . Nature , 2002, 418: 435-438.
[5] Ge Q , Filip L , Bai A ,et al. Inhibition of influenza virus production in virus infected mice by RNA interference[J]. Proc Natl Acad Sci USA, 2004, 101 (23): 8676-8681.
[6] Chen W, Yan W, Du Q,et al. RNA interference targeting VPI inhibits foot and mouth disease virus replication in BHK21 cells and suckling mice[J]. J Virol, 2004, 78 (13): 6900-6907.
[7] Hu WY, Myers CP, Kilzer JM,et al. Inhibition of retroviral pathogenesis by RNA interference[J]. Curr Biol, 2002, 12: 1301-1311.
[8] Jiang M, Milner J. Selective silencing of viral gene expression in HPV2 positive human cervical carcinoma cells treated with siRNAs,a primer of RNA interference[J]. Oncogene, 2002, 21: 6041-6048.
[9] Sui G, Soohoo C, Affar E B, et al. A DNA vector based RNAi technology to suppress gene expression in mammalian cells[J]. Genetics, 2002, 99 (8): 5515-5520.
[10]韩晓荣,柳纪省,杨彬等.亚洲1型口蹄疫病毒反义核酸双向表达载体的构建[J].动物医学进展,2008, 29(3): 5-9.
[11]谢书阳,张敬之,黄淑帧等.应用RNAi抑制细胞中绿色荧光蛋白的表达[J].中华医学遗传学杂志, 2005, 22(4): 431-434.
[12] Coumoul X, Deng CX. RNAi in mice: a promising approach to decipher gene functions in vivo[J]. Biochimie,2006,88(6): 637-643.
[13] JN Leonard, DV Schaffer Antiviral RNAi therapy: emerging approaches for hitting a moving target[J]. Gene Therapy, 2006, 13: 532-540.
[14] Seon-Young Kima, Jae-Ho Leea, Hyun-Seock Shina,et al. The human elongation factor 1 alpha (EF-1α) first intron highly enhances expression of foreign genes from the murine cytomegalovirus promoter[J]. Journal of Biotechnology, 2002, 93(2): 183-187.
[15] Vivek Shukla, Xavier Coumoul, Chu-Xia Deng. RNAi-based conditional gene knockdown in
[1]陈煜,谢小芳. RNAi的作用机制及抗病毒研究进展.世界华人消化杂志, 2006, 14(21): 2123-2129.
[2] Yu JY, DeRuiter SL, Turner DL. RNA interference by expression of short-interfering RNAs and hairpin RNAs in mammalian cells[J]. Proc Natl Acad Sci, 2002, 99(9): 6047-6052.
[3] Novina C.D, and Sharp P.A. The RNAi revolution. Nature 2004; 430: 161-164.
[4] Jacque JM, Triques K , Stevenson M. Modulation of HIV21 replication by RNA interference[J] . Nature , 2002, 418: 435-438.
[5] Ge Q , Filip L , Bai A ,et al. Inhibition of influenza virus production in virus infected mice by RNA interference[J]. Proc Natl Acad Sci USA, 2004, 101 (23): 8676-8681.
[6] Chen W, Yan W, Du Q,et al. RNA interference targeting VPI inhibits foot and mouth disease virus replication in BHK21 cells and suckling mice[J]. J Virol, 2004, 78 (13): 6900-6907.
[7] Hu WY, Myers CP, Kilzer JM,et al. Inhibition of retroviral pathogenesis by RNA interference[J]. Curr Biol, 2002, 12: 1301-1311.
[8] Jiang M, Milner J. Selective silencing of viral gene expression in HPV2 positive human cervical carcinoma cells treated with siRNAs,a primer of RNA interference[J]. Oncogene, 2002, 21: 6041-6048.
[9] Sui G, Soohoo C, Affar E B, et al. A DNA vector based RNAi technology to suppress gene expression in mammalian cells[J]. Genetics, 2002, 99 (8): 5515-5520.
[10]韩晓荣,柳纪省,杨彬等.亚洲1型口蹄疫病毒反义核酸双向表达载体的构建[J].动物医学进展,2008, 29(3): 5-9.
[11]谢书阳,张敬之,黄淑帧等.应用RNAi抑制细胞中绿色荧光蛋白的表达[J].中华医学遗传学杂志, 2005, 22(4): 431-434.
[12] Coumoul X, Deng CX. RNAi in mice: a promising approach to decipher gene functions in vivo[J]. Biochimie,2006,88(6): 637-643.
[13] JN Leonard, DV Schaffer Antiviral RNAi therapy: emerging approaches for hitting a moving target[J]. Gene Therapy, 2006, 13: 532-540.
[14] Seon-Young Kima, Jae-Ho Leea, Hyun-Seock Shina,et al. The human elongation factor 1 alpha (EF-1α) first intron highly enhances expression of foreign genes from the murine cytomegalovirus promoter[J]. Journal of Biotechnology, 2002, 93(2): 183-187.
[15] Vivek Shukla, Xavier Coumoul, Chu-Xia Deng. RNAi-based conditional gene knockdown in
[34] Davenport D, Nichol JAC: Luminescence in Hydromedusae. Proceedings of the Royal Society, Series B 1955, 144:399-411.
[35] Morin JG, Hastings JW: Energy transfer in a bioluminescent system. Journal of Cellular Physiology 1971, 77:313-318.
[36] Anderson JM, Cormier MJ: Lumisomes, the cellular site of bioluminescence in coelenterates. The Journal of Biological Chemistry 1973, 248:2937-2943.
[37] Morise H, Shimomura O, Johnson FH, Winant J: Intermolecular energy transfer in the bioluminecent system of Aequorea. Biochemistry 1974, 13:2656-2662.
[38] Prasher DC, Eckenrode VK, Ward WW, Prendergast FG, Cormier MJ: Primary structure of the Aequorea victoria green fluorescent protein. Gene 1992, 111:229-233.
[39] Bokman SH, Ward WW: Renaturation of Aequorea green fluorescent protein. Biochemical and Biophysical Research Communications 1981, 101:1372-1380.
[40] Cody CW, Prasher DC, Westler WM, Prendergast FG, Ward WW: Chemical-structure of the hexapeptide chromophore of the Aequorea green fluorescent protein. Biochemistry 1993, 32:1212-1218.
[41] Heim R, Prasher DC, Tsien RY: Wavelength mutations and post-translational autoxidation of green fluorescent protein. Proceedings Of the National Academy Of Sciences Of the United States Of America 1994, 91:12501-12504.
[42] Chalfie M, Tu Y, Euskirchen G, Ward WW, Prasher DC: Green fluorescent protein as a marker for gene expression. Science 1994, 263:802-805.
[43] Wang SX, Hazelrigg T: Implications for bcd mRNA localization from spatial distribution of exu protein in Drosophila oogenesis. Nature 1994, 369:400-403.
[44] Baulcombe DC, Chapman S, Cruz SS: Jellyfish green fluorescent protein as a reporter for virus infections. Plant Journal 1995, 7:1045-1053.
[45] Heinlein M, Epel BL, Padgett HS, Beachy RN: Interaction of tobamovirus movement proteins with the plant cytoskeleton. Science 1995, 270:1983-1985.
[46] Haseloff J, Amos B: GFP in plants. Trends in Genetics 1995, 11:328-329.
[47] Hu W, Cheng CL: Expression of Aequorea green fluorescent protein in plant cells. FEBS Letters 1995, 369:331-334.
[48] Sheen J, Hwang SB, Niwa Y, Kobayashi H, Galbraith DW: Green fluorescent protein as a new vital marker in plant cells. Plant Journal 1995, 8:777-78.
[49] Siemering, K.R. Golbik, R., Sever, R. & Haseloff, J. Mutations that supress the thermosensitivity of green fluorescent protein. Current Biology 6:1653-1663, 1996
[50] Haseloff. J. Siemering, K.R., Prasher, D. & Hodge, S. Removal of a cryptic intron and subcellular localisation of green fluorescent protein are required to mark transgenic Arabidopsis plants brightly. Proc. Natl. Acad. Sci. USA, in press.
[51]卢曾军,刘在新.口蹄疫病毒研究进展[J].中国兽医科技, 2003, 33(2):69- 74.
[52]蔡宝祥.家畜传染病学.北京:中国农业出版社,1999. 104 -111.
[53] Carroll A R, Rowlands D J, Carke B E. The complete nucleotide sequence of the RNA coding for the primary translation product of foot-and-mouth disease virus[J].Nucleic Acids Res ,1984 ,12 (5) :2461 -2472.
[54] Forss S, Strebel K, Beck E, et al, Nucleotide sequence and genome organization of Foot-and-mouth disease virus [J]. Nucleic Acids Res, 1984, 12: 6587 - 66011
[55] Pilipenko E V, Blinov V M, Dmitrieva T M. Conservation of the secondary structure elements of 5’-untranslated region cardio and aphthovirus RNAs [J]. Nucleic Acids Research, 1989, 17(14):5701-5711.
[56] Stassinopoulos I A, Belsham G J. A novel protein RNA binding assay: functional interactions of the foot-and-mouth disease virus internal ribosome entry site with cellular proteins [J]. RNA, 2001, 7(1):114-122.
[57] Ohlmann T,Jackson R J.The properties of chimeric picornavirus IRESes show that internal translation initiation sites is influenced by the identity of the IRES and not just the context of the AUG codon[J]. RNA, 1999, 5(6):764-778.
[58] Lopez de Quinto S, Lafuente E, Martinez Salas E. IRES interaction with translation initiation factors: functional characterization of novel RNA contacts with eIF3, eIF4B, and eIF4GII[J]. RNA, 2001, 7(9):1213-1226.
[59] Niepmann M. Effects of potassium and chloride on ribosome association with the RNA of foot-and-mouth disease virus [J]. Virus Res, 2003, 93(1):71-78.
[60]张显升,赵启祖,刘在新,等.口蹄疫病毒基因组内部核糖体进入位点一级和二级结构分析[J].病毒学报, 2002, 18(2):142-148.
[61] Sagedahl A, Giraudo A T, Mello P, et al. Biochemical characterization of an aphthovirus typeC strain Resende attenuated for cattleby serial passages in chicken embryos [J]. Virology, 1987, 157: 366- 374
[62] Gutierrez A, Martinez-Selas E, Pintado B, et al. Specific inhibition of aphthovirus infection by RNAs transcribed from both the 5’and 3’noncoding regions[J].J Viral, 1994, 68 (11): 7436– 7438.
[63] Mateu M G. Antibody recognition of picornaviruses and escap from neutralization: a structural view [J]. Virus Rec, 1995, 38: 1-24.
[64] Wu Q, Moraes M P, Grubman M J. Recombinant adenovirus co-expressing capsid proteins of two serotypes of foot-and-mouth disease virus (FMDV): in vitro characterization and induction of neutralizing antibodies against FMDV in swine [J]. Virus Res, 2003, 93 (2):211-219.
[65] Tosh C, Hemadri D, Sanyal A, et al. Genetic and antigenic analysis of two recently circulating genotypes of type a foot-and-mouth disease virus in India: evidence for positive selection in the capsid coding genes [J]. Arch Virol, 2003, 148(5):853-869.
[66] Balamurugan V, Renji R, Saha S N,et al. Protective immune response of the capsid precursor polypeptide (P1) of foot and mouth disease virus type O produced in Pichia pastoris[J]. Virus Res, 2003, 92(2):141-149.
[67]赵启祖,谢庆阁.口蹄疫病毒抗原位点的研究及应用[A].畜禽重大疫病免疫防制研究进展[C].谢庆阁,翟中和.北京:中国农业出版社,1996,14-23.
[68]徐为燕.兽医病毒学[M].北京:农业出版社,1993:56-59.
[69] Kokuho T, Watanabe S, Yokamizo Y, et al. Production of biologically active,heterodimeric porcine interleukin using a monocistromic baculoviral expression system[J]. Vet Immunol Immunopathol, 1999, 72(3, 4): 289-302.
[70] Gromeier M, Wimmer E, Gorbalenga A E. Genetics, pathogensis and evolution of piconaviru- ses [A]. Domingo E, Webster R, Holland J, et al. Origin and evolution of viruses[C]. Academic Press,1999:287- 343.
[71]江鹏斐,谢庆阁.口蹄疫研究进展[A].谢庆阁,翟中和.兽医生物制品基础研究[C].北京:中国农业科技出版社,1996: 1- 10.
[72] Beard C W, Mason P W. Genetic determinants of altered virulence of Taiwanese foot-and-mouth disease virus [J]. Virol, 2000, 71:987-991.
[73] Knowles N, Davies P R, Henry T, et al. Emergence in Asia of foot-and-mouth disease viruses with altered host range: characterization of alteration in the 3A protein [J]. Virol, 2001, 75(3): 1551-1556.
[74] Wolff J.A., Malone R. W., Williams P, et al. Direct gene transfer into mouse muscle invivo. Science, 1990, 247:1465-1468.
[75]张显升.口蹄疫病毒基因组RNA结构与功能研究进展.病毒学报, 2001, l7(4):375-380.
[76] Grubman MJ, Baxt B.Foot-and-Mouth Disease [J].Clin.Microbiol.Rev. 2004,17:465-493.
[77] DomingoE,EscarmisC,Baranowski E,et al.Evolution of foot-and-mouth disease virus[J].Virus Research, 2003, 91(1):47-63.
[78]农业部畜牧兽医局.一、二、三类动物疫病释义[M].北京:中国农业出版社,2004,1-5.
[79] Guo H, Liu X, Liu Z, et al. Recent outbreaks of foot-and-mouth disease type Asia I in China [J]. J Vet Med B Infect Dis Vet Public Health, 2006, 53(S1): 29-33.
[80]田赠义,尹德华.口蹄疫防疫技术[M].兰州:甘肃民族出版社,2001: 122.
[81]张怀宇.口蹄疫灭活疫苗质量控制和检测技术的研究进展.兽药导刊, 2007: 26-29.
[82]徐全武,申涛.口蹄疫疫苗研究进展[J].新疆畜牧业.2003(3):47-48.
[83]农业部畜牧兽医司.家畜口蹄疫及其防治[M].北京:中国科技出版社,1994:181-184.
[84]薛青红,刘淋涛.口蹄疫病毒3A基因在大肠杆菌中的高效表达[J].微生物学报,2002,5(4): 540 -542.
[85] Bin Yang, Xi Lan, Xueri Li,et al. A novel bi-functional DNA vaccine expressing VP1 protein and producing antisense RNA targeted to 5′UTR of foot-and-mouth disease virus can induce both rapid inhibitory effect and specific immune response in mice[J]. Vaccine, 2008, 26(43), 5477-5483.