LOCUS       sequence-91148-e        8022 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre dependent Cerulean-E2A-SADB19 rabies glycoprotein expression, 
            under EF1a promoter, in AAV virus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8022)
  TITLE     pAAV-FLEX-C-RG
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8022)
  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            "Unpublished"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-FLEX-C-RG
FEATURES             Location/Qualifiers
     source          1..8022
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1441..1450
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1453..1486
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    1549..1582
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(3173..3232)
                     /codon_start=1
                     /product="2A peptide from equine rhinitis A virus 
                     polyprotein"
                     /label=E2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="QCTNYALLKLAGDVESNPGP"
     CDS             complement(join(3248..3958,3959..3961))
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(3290..3311)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3639..3658
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3898..3919
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     protein_bind    complement(3970..4003)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(4074..4107)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4149..4737
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4202..4222)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4620..4631)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4769..5245
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4968..4987)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5285..5425
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5285..5414
                     /label=AAV2 ITR
     rep_origin      5500..5955
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5587..5606)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5797..5818
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5972..5991)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6091..6113
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6151..6169)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6237..6341
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6342..6410,6411..7202)
                     /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(6560..6579)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7373..7961
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7862..7881
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagcc
     1441 gccaccatgg ccataacttc gtataaagta tcctatacga agttatatca aaataggaag
     1501 accaatgctt caccatcgac ccgaattgcc aagcatcacc atcgacccat aacttcgtat
     1561 aatgtatgct atacgaagtt atacttcaac tagttcacag tctggtctcg ccgccgctct
     1621 tgtggctctc ccagctgctg atgatcttgc cgctctgggg ggtcacgctc acctctctgc
     1681 cggtgcctct caggttgtgc tgggtgggct cgcttctgtt cactcttctg cagcaggtca
     1741 tcaggaagat gatcagcatc agggcggtca gggcgccggc gctcagcagc acgtacttgc
     1801 cccagttggg caggcccagg tccacgccgc tcacctggtt gtgcacgtcg ggcaggtgca
     1861 cctccacgaa gtcctcggcc tcgtcgccgt ccttgaacac ggtgctgggg tcggccaggg
     1921 ggtgcaccag ggggatcacg ctgctctcca gcagctccat gtgctgctgc agcaggctgc
     1981 tctgcatctc ggggatcagc acgttgccgt cggggcccag gatgatgccg ttgaagaaca
     2041 cgccgttcac gtgggggtgg catctgccgc ccactctcag gcagcccttg ctgggcagga
     2101 tctcgttcca ggttctcacg ctcttgtagt gggcgtcggc ctccatcagg gtcttgttga
     2161 agatggtgta ggccttgccg aagccgggca ccagctttct caggtggctc agtcttctga
     2221 agctcacgct cttggtggtc atgatgctct ccagggcgtc caggcactcc tctctctttc
     2281 tcaccagctc ctccaccacc aggtgctcga tctcgtcgct tctgaagtcg tgcaggttca
     2341 ccagcttgtc gggggggcac cacttggtct cgttgctggt ctgcatgctc acccaggtgc
     2401 cgtccatcag tctcaggccc agcacgccgc acagcttcag cttgcaggcg cccttcaggc
     2461 tcttgtacag gcctctctcg tccacgaagc cgcaggtctc gctgcccttg ctggctctct
     2521 tgcctctgct gttggtgaag atgtcgcagc tcatgcccag tctggggttc tcgggcatcc
     2581 agatggtgta gtcgtggttg gtgctgcagt aggtgctgct cacggccacg ccgctgcact
     2641 tgccgctggg gaacactctg ctgtgcaggc ttctgtcgta ggggtccagg tcggccacgc
     2701 tggggctgat gatcaccagg ctctccttgg tggtcttcac ggttctcagc catctgtagt
     2761 cggggtaggg gttgtgcagg ctctcctcgt atctggggtc gccggccatc ttccagttgt
     2821 aggcggctct gcaggcgtcg ggggtgggtc tgaagtgctt tctcttgaag gtggtggtca
     2881 cgtagcccac gaagttggtg taggtctcgg cctcggtcac cacgccggtg caggtgaagc
     2941 cgttcacctt gatggccagg atgtagccca ccttcagctc catgtagctg aagccgctca
     3001 ggttggtgca gccctcgtcc tccaccacca ggttgttggg gcagctcagg tggtggatgt
     3061 cgatggggct ccaggggccc agcttgtcgg ggatggtgta gatggggaac ttgccgaagc
     3121 acagggggaa caccagcagg ggcacgaaca gcagggcctg gggcactccg gaaggaccgg
     3181 ggttactttc aacatcgcca gcgagtttca acaaagcgta gttagtacat tgaccactac
     3241 cgctagcctt gtacagctcg tccatgccga gagtgatccc ggcggcggtc acgaactcca
     3301 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagcttggac tgggtgctca
     3361 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt
     3421 ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg gatcttgaag ttggccttga
     3481 tgccgttctt ctgcttgtcg gcggtgatat agacgttgtc gctgatggcg ttgtactcca
     3541 gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat gcccttcagc tcgatgcggt
     3601 tcaccagggt gtcgccctcg aacttcacct cggcgcgggt cttgtagttg ccgtcgtcct
     3661 tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat ggcggacttg aagaagtcgt
     3721 gctgcttcat gtggtcgggg tagcgggcga agcactgcac gccccaggtc agggtggtca
     3781 cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc agggtcagct
     3841 tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg ccgtttacgt
     3901 cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg cccttgctca
     3961 cctcgacgca taacttcgta taggatactt tatacgaagt tatctttggg attcttccta
     4021 ttttgatcca agcatcacca tcgaccctct agtccagatc tcaccatcga cccataactt
     4081 cgtatagcat acattatacg aagttatgtc cctcgaagag gttcgaattc gatatcaagc
     4141 ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     4201 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     4261 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     4321 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     4381 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     4441 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     4501 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
     4561 ggctgctcgc ctatgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     4621 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     4681 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
     4741 gataccgagc gctgctcgag agatctacgg gtggcatccc tgtgacccct ccccagtgcc
     4801 tctcctggcc ctggaagttg ccactccagt gcccaccagc cttgtcctaa taaaattaag
     4861 ttgcatcatt ttgtctgact aggtgtcctt ctataatatt atggggtgga ggggggtggt
     4921 atggagcaag gggcaagttg ggaagacaac ctgtagggcc tgcggggtct attgggaacc
     4981 aagctggagt gcagtggcac aatcttggct cactgcaatc tccgcctcct gggttcaagc
     5041 gattctcctg cctcagcctc ccgagttgtt gggattccag gcatgcatga ccaggctcag
     5101 ctaatttttg tttttttggt agagacgggg tttcaccata ttggccaggc tggtctccaa
     5161 ctcctaatct caggtgatct acccaccttg gcctcccaaa ttgctgggat tacaggcgtg
     5221 aaccactgct cccttccctg tccttctgat tttgtaggta accacgtgcg gaccgagcgg
     5281 ccgcaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact
     5341 gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc
     5401 gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg
     5461 tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat
     5521 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     5581 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     5641 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     5701 ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     5761 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     5821 caacactcaa ccctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg
     5881 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
     5941 taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     6001 gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     6061 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     6121 cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta
     6181 atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg
     6241 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     6301 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     6361 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     6421 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     6481 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     6541 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     6601 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     6661 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     6721 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     6781 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     6841 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     6901 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     6961 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     7021 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     7081 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     7141 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     7201 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     7261 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     7321 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     7381 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     7441 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     7501 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     7561 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     7621 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     7681 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     7741 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     7801 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     7861 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     7921 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     7981 ttttacggtt cctggccttt tgctggcctt ttgctcacat gt
//
