LOCUS       Exported                5847 bp ds-DNA     linear   SYN 05-JUN-2021
DEFINITION  AAV-mediated gene expression from CBh promoter. Contains left AAV2 
            ITR with an 11bp deletion in C region and canonical right AAV2 ITR.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-CBh-mKate2-IRES-MCS (C-gap-WT)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5847)
  TITLE     AAV ITR constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5847)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 5, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5847
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     intron          128..356
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     misc_feature    1131..1683
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1275..1292)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1502..1521
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     misc_feature    1803..2391
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1856..1876)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2274..2285)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2418..2539
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2429..2448)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2483..2502
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2577..2717
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2577..2706
                     /label=AAV2 ITR
     primer_bind     complement(2788..2807)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2907..2929
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2967..2985)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3053..3157
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3158..4018
                     /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(3376..3395)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4189..4777
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4678..4697
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4931..4948
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5065..5086
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5101..5131
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5139..5155
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5144..5166
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5163..5179
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5163..5179
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   5204..5333
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        5431..5734
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg
       61 aaagtttcct tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg
      121 gcgggcggga gtcgctgcga cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc
      181 gccgcccgcc ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc
      241 cttctcctcc gggctgtaat tagctgagca agaggtaagg gtttaaggga tggttggttg
      301 gtggggtatt aatgtttaat tacctggagc acctgcctga aatcactttt tttcaggttg
      361 gaccggtgcc gtttaaacaa caattggaag agctcgtcga cccatggtgt ccgagctgat
      421 caaggagaac atgcacatga agctgtacat ggagggcacc gtgaacaatc accacttcaa
      481 gtgcacatct gagggcgagg gcaagcccta tgagggcacc cagacaatga ggatcaaggc
      541 agtggaggga ggaccactgc ctttcgcctt tgacatcctg gccaccagct tcatgtacgg
      601 ctccaagacc tttatcaacc acacacaggg catccccgac ttctttaagc agagcttccc
      661 tgagggcttt acctgggagc gcgtgaccac atacgaggat ggcggcgtgc tgaccgccac
      721 acaggacaca tccctgcagg atggctgtct gatctataat gtgaagatca ggggcgtgaa
      781 ctttccatct aatggccccg tgatgcagaa gaagaccctg ggatgggagg caagcaccga
      841 gacactgtac ccagcagacg gaggcctgga gggaagggcc gatatggccc tgaagctggt
      901 gggaggagga cacctgatct gcaacctgaa gaccacatat agatctaaga agcctgccaa
      961 gaatctgaag atgccaggcg tgtactatgt ggacaggaga ctggagcgga tcaaggaggc
     1021 cgataaggag acatacgtgg agcagcacga ggtggcagtg gcacggtatt gtgatctgcc
     1081 ctccaagctg ggccaccgct gagaattcga ttacggcggc ccccccccta acgttactgg
     1141 ccgaagccgc ttggaataag gccggtgtgc gtttgtctat atgttatttt ccaccatatt
     1201 gccgtctttt ggcaatgtga gggcccggaa acctggccct gtcttcttga cgagcattcc
     1261 taggggtctt tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg tgaaggaagc
     1321 agttcctctg gaagcttctt gaagacaaac aacgtctgta gcgacccttt gcaggcagcg
     1381 gaacccccca cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat aagatacacc
     1441 tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg atagttgtgg aaagagtcaa
     1501 atggctctcc tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg taccccattg
     1561 tatgggatct gatctggggc ctcggtgcac atgctttaca tgtgtttagt cgaggttaaa
     1621 aaaacgtcta ggccccccga accacgggga cgtggttttc ctttgaaaaa cacgatgata
     1681 atatggccac aaccatgctg gccggcctac ttaattaacc actagttctg gatccttaaa
     1741 actcgaggcc tataacgcgt atagctagct ttgcgatcgc aaaggcgcgc cagcttatcg
     1801 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     1861 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     1921 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     1981 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     2041 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     2101 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     2161 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     2221 tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     2281 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     2341 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     2401 gagcgctgga gagatcttaa gatacattga tgagtttgga caaaccacaa ctagaatgca
     2461 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     2521 aagctgcaat aaacaagttc tgattttgta gttaacacgt gcgaccgagc ggccgcagga
     2581 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     2641 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
     2701 gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     2761 ttcacaccgc atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     2821 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     2881 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     2941 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     3001 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     3061 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     3121 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     3181 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     3241 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     3301 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     3361 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     3421 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     3481 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     3541 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     3601 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     3661 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     3721 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     3781 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     3841 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     3901 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     3961 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     4021 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     4081 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     4141 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     4201 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     4261 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     4321 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     4381 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     4441 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4501 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     4561 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     4621 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     4681 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4741 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4801 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4861 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     4921 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc
     4981 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
     5041 agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc
     5101 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
     5161 cacaggaaac agctatgacc atgattacgc caagcttgca tgccctgcag gcagctgcgc
     5221 gctcgctcgc tcactgaggc cgcccgggcg tcgggcgacc tttggtcgcc cggcctcagt
     5281 gagcgagcga gcgcgcagag agggagtggc caactccatc actaggggtt cctgcgcccg
     5341 cagccgtgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt agcgcgtgcg
     5401 ccaattctgc agacaaattc tagaggtacc cgttacataa cttacggtaa atggcccgcc
     5461 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     5521 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     5581 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     5641 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     5701 gtacatctac gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt
     5761 cactctcccc atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt
     5821 attttgtgca gcgatggggg cgggggg
//
