LOCUS       sequence-373093-        6664 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses nuclear localized tdTomato under CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6664)
  AUTHORS   Feng
  TITLE     Feng Lab AAV plasmid
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6664)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-CAG-2xNLS-tdTomato-WPRE-hGHpolyA
FEATURES             Location/Qualifiers
     source          1..6664
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        860..1163
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1165..1441
                     /label=chicken beta-actin promoter
     primer_bind     1771..1793
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1855..1874
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1917..1926
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1932..1952
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      1959..1968
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1965..3386
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNF
                     EDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLK
                     GEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERS
                     EGRHHLFLYGMDELYK"
     CDS             3393..3413
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    3441..4029
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3494..3514)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3912..3923)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4061..4537
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4260..4279)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4577..4717
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4577..4706
                     /label=AAV2 ITR
     rep_origin      4792..5247
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4879..4898)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5089..5110
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5264..5283)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5383..5405
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5443..5461)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5529..5633
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5634..5702,5703..6494)
                     /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     complement(5852..5871)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggccgcac gcgtaacact agttattaat agtaatcaat tacggggtca ttagttcata
      841 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      901 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      961 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     1021 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     1081 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     1141 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
     1201 tctccccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
     1261 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
     1321 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
     1381 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
     1441 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
     1501 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
     1561 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
     1621 aagccttgag gggctccggg agcgccggca ggaaggaaat gggcggggag ggccttcgtg
     1681 cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg cggggggacg
     1741 gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg accggcggct
     1801 ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc ctgggcaacg
     1861 tgctggttat tgtgctgtct catcattttg gcaaagaatt gcggcccaac ggtaccgcca
     1921 ccatgggaag cccaaagaag aagcgtaagg taggatccgc caccatggtg agcaagggcg
     1981 aggaggtcat caaagagttc atgcgcttca aggtgcgcat ggagggctcc atgaacggcc
     2041 acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca
     2101 agctgaaggt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg tccccccagt
     2161 tcatgtacgg ctccaaggcg tacgtgaagc accccgccga catccccgat tacaagaagc
     2221 tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggtctgg
     2281 tgaccgtgac ccaggactcc tccctgcagg acggcacgct gatctacaag gttaagatgc
     2341 gcggcaccaa cttccccccc gacggccccg taatgcagaa gaagaccatg ggctgggagg
     2401 cctccaccga gcgcctgtac ccccgcgacg gcgtgctgaa gggcgagatc caccaggccc
     2461 tgaagctgaa ggacggcggc cactacctgg tggagttcaa gaccatctac atggccaaga
     2521 agcccgtgca actgcccggc tactactacg tggacaccaa gctggacatc acctcccaca
     2581 acgaggacta caccatcgtg gaacagtacg agcgctccga gggccgccac cacctgttcc
     2641 tggggcatgg caccggcagc accggcagcg gcagctccgg caccgcctcc tccgaggaca
     2701 acaacatggc cgtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga
     2761 acggccacga gttcgagatc gagggcgagg gccgccccta cgagggcacc cagaccgcca
     2821 agctgaaggt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg tccccccagt
     2881 tcatgtacgg ctccaaggcg tacgtgaagc accccgccga catccccgat tacaagaagc
     2941 tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggtctgg
     3001 tgaccgtgac ccaggactcc tccctgcagg acggcacgct gatctacaag gttaagatgc
     3061 gcggcaccaa cttccccccc gacggccccg taatgcagaa gaagaccatg ggctgggagg
     3121 cctccaccga gcgcctgtac ccccgcgacg gcgtgctgaa gggcgagatc caccaggccc
     3181 tgaagctgaa ggacggcggc cactacctgg tggagttcaa gaccatctac atggccaaga
     3241 agcccgtgca actgcccggc tactactacg tggacaccaa gctggacatc acctcccaca
     3301 acgaggacta caccatcgtg gaacagtacg agcgctccga gggccgccac cacctgttcc
     3361 tgtacggcat ggacgagctg tacaagggaa gccccaagaa aaagcggaag gtgtgagaat
     3421 tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg
     3481 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     3541 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     3601 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     3661 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     3721 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     3781 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     3841 cgtcctttcc ttggctgctc gcctatgttg ccacctggat tctgcgcggg acgtccttct
     3901 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     3961 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     4021 cctccccgca tcgataccga gcgctgctcg agagatctac gggtggcatc cctgtgaccc
     4081 ctccccagtg cctctcctgg ccctggaagt tgccactcca gtgcccacca gccttgtcct
     4141 aataaaatta agttgcatca ttttgtctga ctaggtgtcc ttctataata ttatggggtg
     4201 gaggggggtg gtatggagca aggggcaagt tgggaagaca acctgtaggg cctgcggggt
     4261 ctattgggaa ccaagctgga gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc
     4321 ctgggttcaa gcgattctcc tgcctcagcc tcccgagttg ttgggattcc aggcatgcat
     4381 gaccaggctc agctaatttt tgtttttttg gtagagacgg ggtttcacca tattggccag
     4441 gctggtctcc aactcctaat ctcaggtgat ctacccacct tggcctccca aattgctggg
     4501 attacaggcg tgaaccactg ctcccttccc tgtccttctg attttgtagg taaccacgtg
     4561 cggaccgagc ggccgcagga acccctagtg atggagttgg ccactccctc tctgcgcgct
     4621 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
     4681 gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc
     4741 cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct
     4801 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     4861 ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     4921 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     4981 ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct
     5041 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     5101 tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta taagggattt
     5161 tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     5221 ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg
     5281 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     5341 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     5401 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
     5461 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     5521 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     5581 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     5641 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     5701 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     5761 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     5821 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     5881 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     5941 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     6001 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     6061 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     6121 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     6181 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     6241 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     6301 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     6361 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     6421 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     6481 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     6541 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     6601 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     6661 cttc
//
