株单纯疱疹病毒胸苷激酶基因及其应用

摘要

本发明通过PCR方法克隆出高活性的株单纯疱疹病毒1型胸苷激酶(TKc)基因，并测定了其DNA序列，构建了含该基因的逆转录病毒重组表达载体pLTKcSN，并完成了PA317(PLTKcSN)包装细胞株的建立。还应用该包装细胞及其产生的“假型”病毒与GCV配合来治疗恶性肿瘤，在体外的细胞实验及体内的动物试验中，表现其对肿瘤细胞具有杀灭和抑制作用。

主权项

1、一种单纯疱疹病毒1型胸苷激酶基因，其特征在于它是具有中国株单纯疱疹病毒1型胸苷激酶(TKc)基因，其DNA的核苷酸序列及其所编码的氨基酸序列如下：a：TKc基因的氨基酸序列b：TKc基因的核苷酸序列a： 1 Met Ala Ser Tyr Pro Cys His Gln His Ala Ser Ala Phe Asp Gln Alab： 1 ATG GCT TCG TAC CCC TGC CAT CAA CAC GCG TCT GCG TTC GAC CAG GCT 17 Ala Arg Ser Arg Gly His Ser Asn Arg Arg Thr Ala Leu Arg Pro Arg 49 GCG CGT TCT CGC GGC CAT AGC AAC CGA CGT ACG GCG TTG CGC CCT CGC 33 Arg Gln Gln Glu Ala Thr Glu Val Arg Pro Glu Gln Lys Met Pro Thr 97 CGG CAG CAA GAA GCC ACG GAA GTC CGC CCG GAG CAG AAA ATG CCC ACG 49 Leu Leu Arg Val Tyr Ile Asp Gly Pro His Gly Met Gly Lys Thr Thr 145 CTA CTG CGG GTT TAT ATA GAC GGT CCC CAC GGG ATG GGG AAA ACC ACC 65 Thr Thr Gln Leu Leu Val Ala Leu Gly Ser Arg Asp Asp Ile Val Tyr 193 ACC ACG CAA CTG CTG GTG GCC CTG GGT TCG CGC GAC GAT ATC GTC TAC 81 Val Pro Glu Pro Met Thr Tyr Trp Arg Val Leu Gly Ala Ser Glu Thr 241 GTA CCC GAG CCG ATG ACT TAC TGG CGG GTG CTG GGG GCT TCC GAG ACA 97 Ile Ala Asn Ile Tyr Thr Thr Gln His Arg Leu Asp Gln Gly Glu Ile 289 ATC GCG AAC ATC TAC ACC ACA CAA CAC CGC CTC GAC CAG GGT GAG ATA 113 Ser Ala Gly Asp Ala Ala Val Val Met Thr Ser Ala Gln Ile Thr Met 337 TCG GCC GGG GAC GCG GCG GTG GTA ATG ACA AGC GCC CAG ATA ACA ATG 129 Gly Met Pro Tyr Ala Val Thr Asp Ala Val Leu Ala Pro His Ile Gly 385 GGC ATG CCT TAT GCC GTG ACC GAC GCC GTT CTG GCT CCT CAT ATC GGG 145 Gly Glu Ala Gly Ser Ser His Ala Pro Pro Pro Ala Leu Thr Leu Ile 433 GGG GAG GCT GGG AGC TCA CAT GCC CCG CCC CCG GCC CTC ACC CTC ATC 161 Phe Asp Arg His Pro Ile Ala Ala Leu Leu Cys Tyr Pro Ala Ala Arg 481 TTC GAC CGC CAT CCC ATC GCC GCC CTC CTG TGC TAC CCG GCC GCG CGA 177 Tyr Leu Met Gly Ser Met Thr Pro Gln Ala Val Leu Ala Phe Val Ala 529 TAC CTT ATG GGC AGC ATG ACC CCC CAG GCC GTG CTG GCG TTC GTG GCC 193 Leu Ile Pro Pro Thr Leu Pro Gly Thr Asn Ile Val Leu Gly Pro Leu 577 CTC ATC CCG CCG ACC TTG CCC GGC ACC AAC ATC GTG CTT GGG CCC CTT 后续 209 Pro Glu Asp Arg His Ile Asp Arg Leu Ala Lys Arg Gln Arg Pro Gly 625 CCG GAG GAC AGA CAC ATC GAC CGC CTG GCC AAA CGC CAG CGC CCC GGC 225 Glu Arg Leu Asp Leu Ala Met Leu Ala Ala Ile Arg Arg Val Tyr Gly 673 GAG CGG CTG GAC CTG GCT ATG CTG GCT GCG ATT CGC CGC GTT TAC GGG 241 Leu Leu Ala Asn Thr Val Arg Tyr Leu Gln Cys Gly Gly Ser Trp Arg 721 CTA CTT GCC AAT ACG GTG CGG TAT CTG CAG TGC GGC GGG TCG TGG CGG 257 Glu Asp Trp Arg Gln Leu Ser Gly Thr Ala Val Pro Pro Gln Gly Ala 769 GAG GAT TGG AGA CAG CTT TCG GGG ACG GCC GTG CCG CCC CAG GGT GCC 273 Glu Pro Gln Ser Asn Ala Gly Pro Arg Pro His Ile Gly Asp Thr Leu 817 GAG CCC CAG AGC AAC GCG GGC CCA CGA CCC CAT ATC GGG GAC ACG TTA 289 Phe Thr Leu Phe Arg Ala Pro Glu Leu Leu Ala Pro Asn Gly Asp Leu 865 TTT ACC CTG TTT CGG GCC CCC GAG TTG CTG GCC CCC AAC GGC GAC CTG 305 Tyr Asn Val Phe Ala Trp Ala Leu Asp Val Leu Ala Lys Arg Leu Arg 913 TAT AAC GTG TTT GCC TGG GCC TTG GAC GTC TTG GCC AAA CGC CTC CGT 321 Ser Met His Val Phe Ile Leu Asp Tyr Asp Gln Ser Pro Ala Gly Cys 961 TCC ATG CAC GTC TTT ATC CTG GAT TAC GAC CAA TCG CCC GCC GGC TGC 337 Arg Asp Ala Leu Leu Gln Leu Thr Ser Gly Met Val Gln Thr His Val1009 CGG GAC GCC CTG CTG CAA CTT ACC TCC GGG ATG GTC CAG ACC CAC GTC 353 Thr Thr Pro Gly Ser Ile Pro Thr Ile Cys Asp Leu Ala Arg Thr Phe1057 ACC ACC CCA GGC TCC ATA CCG ACG ATC TGC GAC CTG GCG CGC ACG TTT 369 Ala Arg Glu Met Gly Glu Ala Asn ***1105 GCC CGG GAG ATG GGG GAG GCT AAC TGA