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基本信息

質粒圖譜和多克隆位點信息

注意：

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載體描述

Tobacco rattle virus (TRV) is a bipartite, positive-sense RNA virus. TRV1 (i.e., RNA1) encodes 134 and 194 kDa replicase proteins from the genomic RNA, 29 kDa movement protein, and 16 kDa cysteine-rich protein from subgenomic RNAs. TRV1 can replicate and move systemically without TRV2 (i.e., RNA2). TRV2 encodes coat protein from the genomic and two nonstructural proteins from subgenomic RNAs (A). To develop the TRV-based VIGS vector, a cDNA clone of TRV1 or TRV2 (Ppk20 strain) was placed between a duplicated CaMV 35S promoter (2X35S) and nopaline synthase terminator (NOSt) in an Agrobacterium T-DNA vector. In the TRV2 cDNA construct, the nonessential structural genes were replaced with MCS for cloning the target gene sequences. Further, a self-cleaving ribozyme site was included at the 3' end of TRV2. The TRV1 (AF406990)-based viral vector, pTRV1 (linear plasmid 6.791 kb), is used along with pTRV2 for silencing.

The pTRV2 has three variants. The first one is suitable for conventional cloning (AF406991, linear plasmid 9663 nucleotide). The second variant is a gateway-cloning-compatible vector. The third variant is suitable for ligation-independent cloning. Rz, self-cleaving ribozyme; MCS, multiple cloning site; CP, coat protein; MP, movement protein. These vectors were created by Dr. S.P. Dinesh-Kumar, UC Davis, USA. Conventional cloning-compatible (YL156) and ligation-independent-compatible (YY13TRV2) vector plasmid DNA can be obtained from ABRC, Stock No. CD3-1040 vector name YL156. Ligation-independent-compatible TRV2 vector plasmid DNA is ABRC Stock No. CD3-1042, vector name YY13.

載體序列

ORIGIN

        1 ataaaacatt gcacctatgg tgttgccctg gctggggtat gtcagtgatc gcagtagaat

       61 gtactaattg acaagttgga gaatacggta gaacgtcctt atccaacaca gcctttatcc

      121 ctctccctga cgaggttttt gtcagtgtaa tatttctttt tgaactatcc agcttagtac

      181 cgtacgggaa agtgactggt gtgcttatct ttgaaatgtt actttgggtt tcggttcttt

      241 aggttagtaa gaaagcactt gtcttctcat acaaaggaaa acctgagacg tatcgcttac

      301 gaaagtagca atgaaagaaa ggtggtggtt ttaatcgcta ccgcaaaaac gatggggtcg

      361 ttttaattaa cttctcctac gcaagcgtct aaacggacgt tggggttttg ctagtttctt

      421 tagagaaaac tagctaagtc tttaatgtta tcattagaga tggcataaat ataatacttg

      481 tgtctgctga taagatcatt ttaatttgga cgattagact tgttgaacta caggttactg

      541 aatcacttgc gctaatcaac atgggagata tgtacgatga atcatttgac aagtcgggcg

      601 gtcctgctga cttgatggac gattcttggg tggaatcagt ttcgtggaaa gatctgttga

      661 agaagttaca cagcataaaa tttgcactac agtctggtag agatgagatc actgggttac

      721 tagcggcact gaatagacag tgtccttatt caccatatga gcagtttcca gataagaagg

      781 tgtatttcct tttagactca cgggctaaca gtgctcttgg tgtgattcag aacgcttcag

      841 cgttcaagag acgagctgat gagaagaatg cagtggcggg tgttacaaat attcctgcga

      901 atccaaacac aacggttacg acgaaccaag ggagtactac tactaccaag gcgaacactg

      961 gctcgacttt ggaagaagac ttgtacactt attacaaatt cgatgatgcc tctacagctt

     1021 tccacaaatc tctaacttcg ttagagaaca tggagttgaa gagttattac cgaaggaact

     1081 ttgagaaagt attcgggatt aagtttggtg gagcagctgc tagttcatct gcaccgcctc

     1141 cagcgagtgg aggtccgata cgtcctaatc cctagggatt taaggacgtg aactctgttg

     1201 agatctctgt gaaattcaga gggtgggtga taccatattc actgatgcca ttagcgacat

     1261 ctaaataggg ctaattgtga ctaatttgag ggaatttcct ttaccattga cgtcagtgtc

     1321 gttggtagca tttgagtttc gcaatgcacg aattacttag gaagtggctt gacgacacta

     1381 atgtgttatt gttagataat ggtttggtgg tcaaggtacg tagtagagtc ccacatattc

     1441 gcacgtatga agtaattgga aagttgtcag tttttgataa ttcactggga gatgatacgc

     1501 tgtttgaggg aaaagtagag aacgtatttg tttttatgtt caggcggttc ttgtgtgtca

     1561 acaaagatgg acattgttac tcaaggaagc acgatgagct ttattattac ggacgagtgg

     1621 acttagattc tgtgagtaag gttaccgaat tctctagaag gcctccatgg ggatccggta

     1681 ccgagctcac gcgtctcgag gcccgggcat gtcccgaaga cattaaacta cggttcttta

     1741 agtagatccg tgtctgaagt tttaggttca atttaaacct acgagattga cattctcgac

     1801 tgatcttgat tgatcggtaa gtcttttgta atttaatttt ctttttgatt ttattttaaa

     1861 ttgttatctg tttctgtgta tagactgttt gagatcggcg tttggccgac tcattgtctt

     1921 accatagggg aacggacttt gtttgtgttg ttattttatt tgtattttat taaaattctc

     1981 aacgatctga aaaagcctcg cggctaagag attgttgggg ggtgagtaag tacttttaaa

     2041 gtgatgatgg ttacaaaggc aaaaggggta aaacccctcg cctacgtaag cgttattacg

     2101 cccgtctgta cttatatcag tacactgacg agtccctaaa ggacgaaacg ggagaacgct

     2161 agccaccacc accaccacca cgtgtgaatt acaggtgacc agctcgaatt tccccgatcg

     2221 ttcaaacatt tggcaataaa gtttcttaag attgaatcct gttgccggtc ttgcgatgat

     2281 tatcatataa tttctgttga attacgttaa gcatgtaata attaacatgt aatgcatgac

     2341 gttatttatg agatgggttt ttatgattag agtcccgcaa ttatacattt aatacgcgat

     2401 agaaaacaaa atatagcgcg caaactagga taaattatcg cgcgcggtgt catctatgtt

     2461 actagatcgg gaattaaact atcagtgttt gacaggatat attggcgggt aaacctaaga

     2521 gaaaagagcg tttattagaa taacggatat ttaaaagggc gtgaaaaggt ttatccgttc

     2581 gtccatttgt atgtgcatgc caaccacagg gttcccctcg ggatcaaagt actttgatcc

     2641 aacccctccg ctgctatagt gcagtcggct tctgacgttc agtgcagccg tcttctgaaa

     2701 acgacatgtc gcacaagtcc taagttacgc gacaggctgc cgccctgccc ttttcctggc

     2761 gttttcttgt cgcgtgtttt agtcgcataa agtagaatac ttgcgactag aaccggagac

     2821 attacgccat gaacaagagc gccgccgctg gcctgctggg ctatgcccgc gtcagcaccg

     2881 acgaccagga cttgaccaac caacgggccg aactgcacgc ggccggctgc accaagctgt

     2941 tttccgagaa gatcaccggc accaggcgcg accgcccgga gctggccagg atgcttgacc

     3001 acctacgccc tggcgacgtt gtgacagtga ccaggctaga ccgcctggcc cgcagcaccc

     3061 gcgacctact ggacattgcc gagcgcatcc aggaggccgg cgcgggcctg cgtagcctgg

     3121 cagagccgtg ggccgacacc accacgccgg ccggccgcat ggtgttgacc gtgttcgccg

     3181 gcattgccga gttcgagcgt tccctaatca tcgaccgcac ccggagcggg cgcgaggccg

     3241 ccaaggcccg aggcgtgaag tttggccccc gccctaccct caccccggca cagatcgcgc

     3301 acgcccgcga gctgatcgac caggaaggcc gcaccgtgaa agaggcggct gcactgcttg

     3361 gcgtgcatcg ctcgaccctg taccgcgcac ttgagcgcag cgaggaagtg acgcccaccg

     3421 aggccaggcg gcgcggtgcc ttccgtgagg acgcattgac cgaggccgac gccctggcgg

     3481 ccgccgagaa tgaacgccaa gaggaacaag catgaaaccg caccaggacg gccaggacga

     3541 accgtttttc attaccgaag agatcgaggc ggagatgatc gcggccgggt acgtgttcga

     3601 gccgcccgcg cacgtctcaa ccgtgcggct gcatgaaatc ctggccggtt tgtctgatgc

     3661 caagctggcg gcctggccgg ccagcttggc cgctgaagaa accgagcgcc gccgtctaaa

     3721 aaggtgatgt gtatttgagt aaaacagctt gcgtcatgcg gtcgctgcgt atatgatgcg

     3781 atgagtaaat aaacaaatac gcaaggggaa cgcatgaagg ttatcgctgt acttaaccag

     3841 aaaggcgggt caggcaagac gaccatcgca acccatctag cccgcgccct gcaactcgcc

     3901 ggggccgatg ttctgttagt cgattccgat ccccagggca gtgcccgcga ttgggcggcc

     3961 gtgcgggaag atcaaccgct aaccgttgtc ggcatcgacc gcccgacgat tgaccgcgac

     4021 gtgaaggcca tcggccggcg cgacttcgta gtgatcgacg gagcgcccca ggcggcggac

     4081 ttggctgtgt ccgcgatcaa ggcagccgac ttcgtgctga ttccggtgca gccaagccct

     4141 tacgacatat gggccaccgc cgacctggtg gagctggtta agcagcgcat tgaggtcacg

     4201 gatggaaggc tacaagcggc ctttgtcgtg tcgcgggcga tcaaaggcac gcgcatcggc

     4261 ggtgaggttg ccgaggcgct ggccgggtac gagctgccca ttcttgagtc ccgtatcacg

     4321 cagcgcgtga gctacccagg cactgccgcc gccggcacaa ccgttcttga atcagaaccc

     4381 gagggcgacg ctgcccgcga ggtccaggcg ctggccgctg aaattaaatc aaaactcatt

     4441 tgagttaatg aggtaaagag aaaatgagca aaagcacaaa cacgctaagt gccggccgtc

     4501 cgagcgcacg cagcagcaag gctgcaacgt tggccagcct ggcagacacg ccagccatga

     4561 agcgggtcaa ctttcagttg ccggcggagg atcacaccaa gctgaagatg tacgcggtac

     4621 gccaaggcaa gaccattacc gagctgctat ctgaatacat cgcgcagcta ccagagtaaa

     4681 tgagcaaatg aataaatgag tagatgaatt ttagcggcta aaggaggcgg catggaaaat

     4741 caagaacaac caggcaccga cgccgtggaa tgccccatgt gtggaggaac gggcggttgg

     4801 ccaggcgtaa gcggctgggt tgtctgccgg ccctgcaatg gcactggaac ccccaagccc

     4861 gaggaatcgg cgtgacggtc gcaaaccatc cggcccggta caaatcggcg cggcgctggg

     4921 tgatgacctg gtggagaagt tgaaggccgc gcaggccgcc cagcggcaac gcatcgaggc

     4981 agaagcacgc cccggtgaat cgtggcaagc ggccgctgat cgaatccgca aagaatcccg

     5041 gcaaccgccg gcagccggtg cgccgtcgat taggaagccg cccaagggcg acgagcaacc

     5101 agattttttc gttccgatgc tctatgacgt gggcacccgc gatagtcgca gcatcatgga

     5161 cgtggccgtt ttccgtctgt cgaagcgtga ccgacgagct ggcgaggtga tccgctacga

     5221 gcttccagac gggcacgtag aggtttccgc agggccggcc ggcatggcca gtgtgtggga

     5281 ttacgacctg gtactgatgg cggtttccca tctaaccgaa tccatgaacc gataccggga

     5341 agggaaggga gacaagcccg gccgcgtgtt ccgtccacac gttgcggacg tactcaagtt

     5401 ctgccggcga gccgatggcg gaaagcagaa agacgacctg gtagaaacct gcattcggtt

     5461 aaacaccacg cacgttgcca tgcagcgtac gaagaaggcc aagaacggcc gcctggtgac

     5521 ggtatccgag ggtgaagcct tgattagccg ctacaagatc gtaaagagcg aaaccgggcg

     5581 gccggagtac atcgagatcg agctagctga ttggatgtac cgcgagatca cagaaggcaa

     5641 gaacccggac gtgctgacgg ttcaccccga ttactttttg atcgatcccg gcatcggccg

     5701 ttttctctac cgcctggcac gccgcgccgc aggcaaggca gaagccagat ggttgttcaa

     5761 gacgatctac gaacgcagtg gcagcgccgg agagttcaag aagttctgtt tcaccgtgcg

     5821 caagctgatc gggtcaaatg acctgccgga gtacgatttg aaggaggagg cggggcaggc

     5881 tggcccgatc ctagtcatgc gctaccgcaa cctgatcgag ggcgaagcat ccgccggttc

     5941 ctaatgtacg gagcagatgc tagggcaaat tgccctagca ggggaaaaag gtcgaaaagg

     6001 tctctttcct gtggatagca cgtacattgg gaacccaaag ccgtacattg ggaaccggaa

     6061 cccgtacatt gggaacccaa agccgtacat tgggaaccgg tcacacatgt aagtgactga

     6121 tataaaagag aaaaaaggcg atttttccgc ctaaaactct ttaaaactta ttaaaactct

     6181 taaaacccgc ctggcctgtg cataactgtc tggccagcgc acagccgaag agctgcaaaa

     6241 agcgcctacc cttcggtcgc tgcgctccct acgccccgcc gcttcgcgtc ggcctatcgc

     6301 ggccgctggc cgctcaaaaa tggctggcct acggccaggc aatctaccag ggcgcggaca

     6361 agccgcgccg tcgccactcg accgccggcg cccacatcaa ggcaccctgc ctcgcgcgtt

     6421 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc

     6481 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt

     6541 gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta

     6601 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag

     6661 atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct

     6721 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt

     6781 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc

     6841 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga

     6901 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata

     6961 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac

     7021 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg

     7081 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc

     7141 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag

     7201 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt

     7261 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt

     7321 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg

     7381 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac

     7441 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca

     7501 gtggaacgaa aactcacgtt aagggatttt ggtcatgcat tctaggtact aaaacaattc

     7561 atccagtaaa atataatatt ttattttctc ccaatcaggc ttgatcccca gtaagtcaaa

     7621 aaatagctcg acatactgtt cttccccgat atcctccctg atcgaccgga cgcagaaggc

     7681 aatgtcatac cacttgtccg ccctgccgct tctcccaaga tcaataaagc cacttacttt

     7741 gccatctttc acaaagatgt tgctgtctcc caggtcgccg tgggaaaaga caagttcctc

     7801 ttcgggcttt tccgtcttta aaaaatcata cagctcgcgc ggatctttaa atggagtgtc

     7861 ttcttcccag ttttcgcaat ccacatcggc cagatcgtta ttcagtaagt aatccaattc

     7921 ggctaagcgg ctgtctaagc tattcgtata gggacaatcc gatatgtcga tggagtgaaa

     7981 gagcctgatg cactccgcat acagctcgat aatcttttca gggctttgtt catcttcata

     8041 ctcttccgag caaaggacgc catcggcctc actcatgagc agattgctcc agccatcatg

     8101 ccgttcaaag tgcaggacct ttggaacagg cagctttcct tccagccata gcatcatgtc

     8161 cttttcccgt tccacatcat aggtggtccc tttataccgg ctgtccgtca tttttaaata

     8221 taggttttca ttttctccca ccagcttata taccttagca ggagacattc cttccgtatc

     8281 ttttacgcag cggtattttt cgatcagttt tttcaattcc ggtgatattc tcattttagc

     8341 catttattat ttccttcctc ttttctacag tatttaaaga taccccaaga agctaattat

     8401 aacaagacga actccaattc actgttcctt gcattctaaa accttaaata ccagaaaaca

     8461 gctttttcaa agttgttttc aaagttggcg tataacatag tatcgacgga gccgattttg

     8521 aaaccgcggt gatcacaggc agcaacgctc tgtcatcgtt acaatcaaca tgctaccctc

     8581 cgcgagatca tccgtgtttc aaacccggca gcttagttgc cgttcttccg aatagcatcg

     8641 gtaacatgag caaagtctgc cgccttacaa cggctctccc gctgacgccg tcccggactg

     8701 atgggctgcc tgtatcgagt ggtgattttg tgccgagctg ccggtcgggg agctgttggc

     8761 tggctggtgg caggatatat tgtggtgtaa acaaattgac gcttagacaa cttaataaca

     8821 cattgcggac gtttttaatg tactgaatta acgccgaatt aattcctagg ccaccatgtt

     8881 gggcccggcg cgccaagctt gcatgcctgc aggtcaacat ggtggagcac gacactctcg

     8941 tctactccaa gaatatcaaa gatacagtct cagaagacca gagggctatt gagacttttc

     9001 aacaaagggt aatatcggga aacctcctcg gattccattg cccagctatc tgtcacttca

     9061 tcgaaaggac agtagaaaag gaagatggct tctacaaatg ccatcattgc gataaaggaa

     9121 aggctatcgt tcaagatgcc tctaccgaca gtggtcccaa agatggaccc ccacccacga

     9181 ggaacatcgt ggaaaaagaa gacgttccaa ccacgtcttc aaagcaagtg gattgatgtg

     9241 atggtcaaca tggtggagca cgacactctc gtctactcca agaatatcaa agatacagtc

     9301 tcagaagacc agagggctat tgagactttt caacaaaggg taatatcggg aaacctcctc

     9361 ggattccatt gcccagctat ctgtcacttc atcgaaagga cagtagaaaa ggaagatggc

     9421 ttctacaaat gccatcattg cgataaagga aaggctatcg ttcaagatgc ctctaccgac

     9481 agtggtccca aagatggacc cccacccacg aggaacatcg tggaaaaaga agacgttcca

     9541 accacgtctt caaagcaagt ggattgatgt gatatctcca ctgacgtaag ggatgacgca

     9601 caatcccact atccttcgca agacccttcc tctatataag gaagttcatt tcatttggag

     9661 agg

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