LOCUS       sequence-358488-        7070 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express hM4Di in cells not expressing Cre..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7070)
  AUTHORS   Beier KT, Kim CK, Hoerbelt P, Hung LW, Heifets BD, DeLoach KE, Mosca
            TJ, Neuner S, Deisseroth K, Luo L, Malenka RC
  TITLE     Rabies screen reveals GPe control of cocaine-triggered plasticity.
  JOURNAL   Nature. 2017 Sep 21;549(7672):345-350. doi: 10.1038/nature23888. 
            Epub 2017 Sep 13.
  PUBMED    28902833
REFERENCE   2  (bases 1 to 7070)
  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"; doi: 
            "10.1038/nature23888"; journalName: "Nature"; date: "2017-09-21- 
            21"; volume: "549"; issue: "7672"; pages: "345-350"
COMMENT     Sequence Label: pAAV hSyn FLEx-OFF hM4Di-mCherry
FEATURES             Location/Qualifiers
     source          1..7070
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..448
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    524..557
                     /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    620..653
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      2112..2121
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2118..2828
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2263..2281)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2531..2550
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2838..2871)
                     /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(2942..2975)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3023..3611
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3076..3096)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3494..3505)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3643..4119
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3842..3861)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4159..4299
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4159..4288
                     /label=AAV2 ITR
     rep_origin      4374..4829
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4461..4480)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4671..4692
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4846..4865)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4965..4987
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5025..5043)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5111..5215
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5216..5284,5285..6076)
                     /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(5434..5453)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6247..6835
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6736..6755
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   6897..7026
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 agtgcaagtg ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga
       61 ccccgaccca ctggacaagc acccaacccc cattccccaa attgcgcatc ccctatcaga
      121 gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg
      181 gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc
      241 gctgacgtca ctcgccggtc ccccgcaaac tccccttccc ggccaccttg gtcgcgtccg
      301 cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc gagatagggg ggcacgggcg
      361 cgaccatctg cgctgcggcg ccggcgactc agcgctgcct cagtctgcgg tgggcagcgg
      421 aggagtcgtg tcgtgcctga gagcgcagtc gagaaggtac cggatcctgt gctgtctcat
      481 cattttggca aagaattgat ttgataccgc gggcccggcc gggataactt cgtataaagt
      541 atcctatacg aagttatatc aaaataggaa gaccaatgct tcaccatcga cccgaattgc
      601 caagcatcac catcgacaca taacttcgta taatgtatgc tatacgaagt tatgtcgaca
      661 tggccaactt cacacctgtc aatggcagct cgggcaatca gtccgtgcgc ctggtcacgt
      721 catcatccca caatcgctat gagacggtgg aaatggtctt cattgccaca gtgacaggct
      781 ccctgagcct ggtgactgtc gtgggcaaca tcctggtgat gctgtccatc aaggtcaaca
      841 ggcagctgca gacagtcaac aactacttcc tcttcagcct ggcgtgtgct gatctcatca
      901 taggcgcctt ctccatgaac ctctacaccg tgtacatcat caagggctac tggcccctgg
      961 gcgccgtggt ctgcgacctg tggctggccc tggactgcgt ggtgagcaac gcctccgtca
     1021 tgaaccttct catcatcagc tttgaccgct acttctgcgt caccaagcct ctcacctacc
     1081 ctgcccggcg caccaccaag atggcaggcc tcatgattgc tgctgcctgg gtactgtcct
     1141 tcgtgctctg ggcgcctgcc atcttgttct ggcagtttgt ggtgggtaag cggacggtgc
     1201 ccgacaacca gtgcttcatc cagttcctgt ccaacccagc agtgaccttt ggcacagcca
     1261 ttgctggctt ctacctgcct gtggtcatca tgacggtgct gtacatccac atctccctgg
     1321 ccagtcgcag ccgagtccac aagcaccggc ccgagggccc gaaggagaag aaagccaaga
     1381 cgctggcctt cctcaagagc ccactaatga agcagagcgt caagaagccc ccgcccgggg
     1441 aggccgcccg ggaggagctg cgcaatggca agctggagga ggcccccccg ccagcgctgc
     1501 caccgccacc gcgccccgtg gctgataagg acacttccaa tgagtccagc tcaggcagtg
     1561 ccacccagaa caccaaggaa cgcccagcca cagagctgtc caccacagag gccaccacgc
     1621 ccgccatgcc cgcccctccc ctgcagccgc gggccctcaa cccagcctcc agatggtcca
     1681 agatccagat tgtgacgaag cagacaggca atgagtgtgt gacagccatt gagattgtgc
     1741 ctgccacgcc ggctggcatg cgccctgcgg ccaacgtggc ccgcaagttc gccagcatcg
     1801 ctcgcaacca ggtgcgcaag aagcggcaga tggcggcccg ggagcgcaaa gtgacacgaa
     1861 cgatctttgc cattctgctg gccttcatcc tcacctggac gccctacaac gtcatggtcc
     1921 tggtgaacac cttctgccag agctgcatcc ctgacacggt gtggtccatt ggctactggc
     1981 tctgctacgt caacagcacc atcaaccctg cctgctatgc tctgtgcaac gccaccttta
     2041 aaaagacctt ccggcacctg ctgctgtgcc agtatcggaa catcggcact gccaggcggg
     2101 atccaccggt cgccaccatg gtgagcaagg gcgaggagga taacatggcc atcatcaagg
     2161 agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg
     2221 agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca
     2281 agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca
     2341 aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg
     2401 gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg
     2461 actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc
     2521 cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga
     2581 tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg
     2641 gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc
     2701 ccggcgccta caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca
     2761 tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt
     2821 acaagtaagg cgcgcctata acttcgtata ggatacttta tacgaagtta tcattgggat
     2881 tcttcctatt ttgatccaag catcaccatc gaccctctag tccagatctc accatcgacc
     2941 cataacttcg tatagcatac attatacgaa gttataagct tatatgcggc cgatcgatga
     3001 attcgatatc aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac
     3061 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     3121 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     3181 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     3241 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     3301 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     3361 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc
     3421 atcgtccttt ccttggctgc tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt
     3481 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     3541 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     3601 cgcctccccg catcgatacc gagcgctgct cgagagatct acgggtggca tccctgtgac
     3661 ccctccccag tgcctctcct ggccctggaa gttgccactc cagtgcccac cagccttgtc
     3721 ctaataaaat taagttgcat cattttgtct gactaggtgt ccttctataa tattatgggg
     3781 tggagggggg tggtatggag caaggggcaa gttgggaaga caacctgtag ggcctgcggg
     3841 gtctattggg aaccaagctg gagtgcagtg gcacaatctt ggctcactgc aatctccgcc
     3901 tcctgggttc aagcgattct cctgcctcag cctcccgagt tgttgggatt ccaggcatgc
     3961 atgaccaggc tcagctaatt tttgtttttt tggtagagac ggggtttcac catattggcc
     4021 aggctggtct ccaactccta atctcaggtg atctacccac cttggcctcc caaattgctg
     4081 ggattacagg cgtgaaccac tgctcccttc cctgtccttc tgattttgta ggtaaccacg
     4141 tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
     4201 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
     4261 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc
     4321 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
     4381 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     4441 tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     4501 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     4561 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     4621 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     4681 gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat
     4741 tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     4801 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
     4861 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     4921 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     4981 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     5041 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     5101 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     5161 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     5221 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     5281 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     5341 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     5401 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     5461 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     5521 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     5581 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     5641 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     5701 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     5761 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     5821 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     5881 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     5941 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     6001 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     6061 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     6121 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     6181 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     6241 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     6301 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     6361 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     6421 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     6481 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     6541 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     6601 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     6661 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     6721 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     6781 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     6841 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtcctg
     6901 caggcagctg cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg acctttggtc
     6961 gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg
     7021 gttcctgcgg ccgcacgcgt gtgtctagac tgcagagggc cctgcgtatg
//
