LOCUS       pCAG-D12L               6577 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Express vaccinia virus D12L in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pCAG-D12L
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6577)
  AUTHORS   Kanai Y, Komoto S, Kawagishi T, Nouda R, Nagasawa N, Onishi M, 
            Matsuura Y, Taniguchi K, Kobayashi T
  TITLE     Entirely plasmid-based reverse genetics system for rotaviruses.
  JOURNAL   Proc Natl Acad Sci U S A. 2017 Jan 30. pii: 201618424. doi: 
            10.1073/pnas.1618424114.
  PUBMED    28137864
REFERENCE   2  (bases 1 to 6577)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            //www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6577
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..657
                     /label=chicken beta-actin promoter
     intron          658..1672
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1596..1618
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1680..1699
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(2722..2741)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2787..2842
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2841..2860)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3194..3214)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     3441..3460
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3621..4220
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3621..3640)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4117..4137
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4450..4584
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4487..4506)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4541..4560
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4652..4669)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4823..5411)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4903..4922)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5582..6442)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6205..6224
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6443..6547)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct
      421 cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga
      481 tgggggcggg gggggggggg gggggcgcca ggcggggcgg ggcggggcga ggggcggggc
      541 ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct
      601 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga
      661 gtcgctgcgt tgccttcgcc ccgtgccccg ctccgcgccg cctcgcgccg cccgccccgg
      721 ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc
      781 tgtaattagc gcttggttta atgacggctc gtttcttttc tgtggctgcg tgaaagcctt
      841 aaagggctcc gggagggccc tttgtgcggg ggggagcggc tcggggggtg cgtgcgtgtg
      901 tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg agcgctgcgg
      961 gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg ccgggggcgg
     1021 tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg gtgtgtgcgt
     1081 gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg caaccccccc tgcacccccc
     1141 tccccgagtt gctgagcacg gcccggcttc gggtgcgggg ctccgtacgg ggcgtggcgc
     1201 ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg gtgccgggcg gggcggggcc
     1261 gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg ccggcggctg
     1321 tcgaggcgcg gcgagccgca accattgcct tttatggtaa tcgtgcgaga gggcgcaagg
     1381 acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg gggagggcct
     1501 tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct gtccgcgggg
     1561 ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc gtgtgaccgg
     1621 cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac agctcctggg
     1681 caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattaatta agtttaaacg
     1741 gcgcgccgat atcttaagat ctatggatga aattgtaaaa aatatccggg agggaacgca
     1801 tgtccttctt ccattttatg aaacattgcc agaacttaat ctgtctctag gtaaaagccc
     1861 attacctagt ctggaatacg gagctaatta ctttcttcag atttctagag ttaatgatct
     1921 aaatagaatg ccgaccgaca tgttaaaact ttttacacat gatatcatgt taccagaaag
     1981 cgatctagat aaagtctatg aaattttaaa gattaatagc gtaaagtatt atgggaggag
     2041 tactaaagcg gacgccgtag ttgccgacct cagcgcacgc aataaactgt tcaaacgtga
     2101 acgagatgct attaaatcta ataatcatct cactgaaaac aatctataca ttagcgatta
     2161 taagatgtta accttcgacg tgtttcgacc attatttgat tttgtaaacg aaaaatattg
     2221 tattattaaa cttccaactt tattcggtag aggtgtaatc gatactatga gaatatattg
     2281 tagtctcttt aaaaatgtta gactgctaaa atgcgtaagc gatagctggt tgaaagatag
     2341 cgccattatg gtggctagtg atgtttgtaa aaaaaatttg gatttattta tgtctcatgt
     2401 taagtccgtc actaagtctt cttcttggaa ggatgtgaac agtgttcaat ttagtatttt
     2461 aaacaatcca gtggatacgg aattcattaa taagttctta gagttttcga atagagtata
     2521 cgaagctctc tattacgttc actcgttgct ttattctagt atgacttctg attcaaaaag
     2581 tatcgaaaac aaacatcaga gaagactagt taaactactg ctgtgaagat cttcgaattc
     2641 actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa tgccctggct
     2701 cacaaatacc actgagatcg atctttttcc ctctgccaaa aattatgggg acatcatgaa
     2761 gccccttgag catctgactt ctggctaata aaggaaattt attttcattg caatagtgtg
     2821 ttggaatttt ttgtgtctct cactcggaag gacatatggg agggcaaatc atttaaaaca
     2881 tcagaatgag tatttggttt agagtttggc aacatatgcc catatgctgg ctgccatgaa
     2941 caaaggttgg ctataaagag gtcatcagta tatgaaacag ccccctgctg tccattcctt
     3001 attccataga aaagccttga cttgaggtta gatttttttt atattttgtt ttgtgttatt
     3061 tttttcttta acatccctaa aattttcctt acatgtttta ctagccagat ttttcctcct
     3121 ctcctgacta ctcccagtca tagctgtccc tcttctctta tggagatccc tcgacctgca
     3181 gcccaagctc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3241 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3301 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3361 ccgcccctaa ctccgcccat cccgccccta actccgccca gtttccgccc ttctccgccc
     3421 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     3481 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gcttgcatgc
     3541 ctgcaggtcg gccgccacga ccggtgccgc caccatcccc tgacccacgc ccctgacccc
     3601 tcacaaggag acgaccttcc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     3661 acgacgtccc ccgggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     3721 gccacaccgt cgacccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     3781 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     3841 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     3901 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     3961 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     4021 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     4081 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     4141 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     4201 cccgcaagcc cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg
     4261 cacgacccca tggctccgac cgaagccacc cggggcggcc ccgccgaccc cgcacccgcc
     4321 cccgaggccc accgactcta gaggatcata atcagccata ccacatttgt agaggtttta
     4381 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
     4441 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     4501 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     4561 aatgtatctt atcatgtctg gatccgctgc attaatgaat cggccaacgc gcggggagag
     4621 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4681 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4741 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4801 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4861 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4921 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4981 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     5041 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     5101 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     5161 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     5221 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     5281 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     5341 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5401 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5461 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5521 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5581 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5641 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5701 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5761 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5821 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5881 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5941 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6001 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6061 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     6121 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6181 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6241 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     6301 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6361 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6421 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6481 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6541 ttccgcgcac atttccccga aaagtgccac ctgggtc
//