LOCUS       sequence-362556-        7498 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent chemogenetic inhibitor expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7498)
  AUTHORS   Magnus CJ, Lee PH, Bonaventura J, Zemla R, Gomez JL, Ramirez MH, Hu 
            X, Galvan A, Basu J, Michaelides M, Sternson SM
  TITLE     Ultrapotent chemogenetics for research and potential clinical 
            applications.
  JOURNAL   Science. 2019 Mar 14. pii: science.aav5282. doi: 
            10.1126/science.aav5282.
  PUBMED    30872534
REFERENCE   2  (bases 1 to 7498)
  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: "Science. 
            2019 Mar 14. pii: science.aav5282. doi: 10.1126/science.aav5282."
COMMENT     Sequence Label: AAV SYN flex PSAM4 GlyR IRES EGFP
FEATURES             Location/Qualifiers
     source          1..7498
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   56..185
                     /label=AAV2 ITR
     polyA_signal    347..468
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(384..403)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     438..457
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    545..1133
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             651..662
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1060..1080
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    1201..1234
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1305..1338
                     /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)."
     misc_feature    join(2703..3276,3277..3279,3280..3288)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2870..2887)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             join(3301..3303,3304..4017)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(3343..3364)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3604..3623)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3951..3972
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4030..4063)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(4126..4159)
                     /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)."
     primer_bind     complement(4178..4197)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          4180..4312
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     repeat_region   4911..5040
                     /label=AAV2 ITR
     rep_origin      5112..5567
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5199..5218)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5409..5430
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5593..5697
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5698..5766,5767..6558)
                     /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(5916..5935)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6729..7317
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7218..7237
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctgcg
       61 cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg
      121 cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg
      181 ttccttgtag ttaatgatta acccgccatg ctacttatct acgtagccat gctctaggaa
      241 gatccttatc gattttacca catttgtaga ggttttactt gctttaaaaa acctcccaca
      301 tctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc
      361 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
      421 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgctc
      481 gaagcggccg gccgcccggg cgacctagat gcattcgacg gccgcccggg tcgacggtat
      541 cgatgcgggg aggcggccca aagggagatc cgactcgtct gagggcgaag gcgaagacgc
      601 ggaagaggcc gcagagccgg cagcaggccg cgggaaggaa ggtccgctgg attgagggcc
      661 gaagggacgt agcagaagga cgtcccgcgc agaatccagg tggcaacaca ggcgagcagc
      721 caaggaaagg acgatgattt ccccgacaac accacggaat tgtcagtgcc caacagccga
      781 gcccctgtcc agcagcgggc aaggcaggcg gcgatgagtt ccgccgtggc aatagggagg
      841 gggaaagcga aagtcccgga aaggagctga caggtggtgg caatgcccca accagtgggg
      901 gttgcgtcag caaacacagt gcacaccacg ccacgttgcc tgacaacggg ccacaactcc
      961 tcataaagag acagcaacca ggatttatac aaggaggaga aaatgaaagc catacgggaa
     1021 gcaatagcat gatacaaagg cattaaagca gcgtatccac atagcgtaaa aggagcaaca
     1081 tagttaagaa taccagtcaa tctttcacaa attttgtaat ccagaggttg attcttaaga
     1141 gcttcgagtg gatcatctgc aaattcgccc ttttcagtgt ttcgaacctc ttcgagggac
     1201 ataacttcgt ataatgtatg ctatacgaag ttatgggtcg atggtgagat ctggactaga
     1261 gggtcgatgg tgatgcttgg atcaaaatag gaagaatccc aatgataact tcgtataaag
     1321 tatcctatac gaagttatac tagtcatatg ggatccgaat tcgctagcgc caccatgcgc
     1381 tgttctccag gcggcgtgtg gctcgccctg gctgcttccc ttctgcacgt tagcctgcag
     1441 ggtgagttcc agcgcaaact gtataaggag cttgttaaga attataaccc cctggagcgg
     1501 ccggtcgcaa atgattccca gccactgaca gtgtacttca gcctctcctt gctgcagatc
     1561 atggacgtgg atgaaaagaa ccaggtgctg accactaata tttggttgca gatgtcctgg
     1621 accgatcact acttgcagtg gaatgtgagc gaatacccag gtgtaaagac tgtaagattc
     1681 cctgacggcc aaatctggaa accagatatc ctgctgtaca acagcgcaga cgaaaggttt
     1741 gatgcaacat ttcacaccaa cgtgggagtc aattcttcag gccactgcct gtacctgccc
     1801 cctggaatct tcaagtcctc atgctatatc gacgtccgct ggtttccctt cgacgtccag
     1861 cactgcaaac tcaaattcgg gagctggagc tacggcggat ggagcctgga tctgcaaatg
     1921 caggaggctg acatctctgg ttacatcccg aatggggagt gggaccttgt gggaatcccc
     1981 ggtaaaagaa gcgagcgatt ttatgaatgc tgcaaagagc ccttcccaga tgtcaccttc
     2041 acagtgacca tgcggagacg catgggttat tatctgatcc aaatgtatat cccaagcttg
     2101 cttatagtga ttttgtcatg gatctccttc tggattaata tggacgccgc tccagctagg
     2161 gtcggactgg gcatcaccac agtgctgaca atgactactc agagctcagg cagccgagcc
     2221 agcttgccca aggtttctta cgtgaaggcc atcgatatct ggatggctgt ctgccttctg
     2281 tttgtcttca gcgcactgct ggaatacgcc gctgtcaatt ttgtgtctcg acagcataaa
     2341 gagctgttgc ggttcagaag aaaacgacgc caccacaaag aggatgaggc aggagaagga
     2401 cgcttcaact ttagcgccta tggtatggga cctgcttgcc tccaggctaa agacggaatt
     2461 tccgtgaagg gagccaacaa tagcaacaca accaacccac cccctgctcc atctaagagc
     2521 ccggaggaaa tgcgcaaact ctttattcag agagcgaaaa agatcgacaa aatctcccgg
     2581 atcggattcc ccatggcttt cctgattttc aacatgtttt attggatcat ctacaagatt
     2641 gtgcgaaggg aggacgtaca caaccagtaa gcggccgcaa ttccccccgc cccccccccc
     2701 cccccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga ataaggccgg
     2761 tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa tgtgagggcc
     2821 cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc tctcgccaaa
     2881 ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga
     2941 caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg cgacaggtgc
     3001 ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca accccagtgc
     3061 cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctaagc gtattcaaca
     3121 aggggctgaa ggatgcccag aaggtacccc attgtatggg atctgatctg gggcctcggt
     3181 gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg
     3241 gggacgtggt tttcctttga aaaacacgat gataatatgg ccacaaccat gggggatccg
     3301 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     3361 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     3421 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     3481 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     3541 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     3601 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     3661 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     3721 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     3781 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     3841 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     3901 ctgagcaccc agtccaagct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     3961 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaatct
     4021 agcaagctta taacttcgta tagcatacat tatacgaagt tatgtgtcga tggtgatgct
     4081 tggcaattcg ggtcgatggt gaagcattgg tcttcctatt ttgatataac ttcgtatagg
     4141 atactttata cgaagttatg gggggcgcgc catcatcacc tgtggagaga aaggcaaagt
     4201 ggatgtcagt aagaccaata ggtgcctatc agaaacgcaa gagtcttctc tgtctcgaca
     4261 agcccagttt ctattggtct ccttaaacct gtcttgtaac cttgatactt acctgatcca
     4321 acggctgcga cttggggcag ggggtcctag gggtggtctg gccaggagcc gcgggggcgg
     4381 actgcgcggt gcccaggagc acagctgcgc tctcaggcac gacacgactc ctccgctgcc
     4441 caccgcagac tgaggcagcg ctgagtcgcc ggcgccgcag cgcagatggt cgcgcccgtg
     4501 cccccctatc tcgcgcctcg cgtggtgcgg tccggctggg ccggcggcgg cgcggacgcg
     4561 accaaggtgg ccgggaaggg gagtttgcgg gggaccggcg agtgacgtca gcgcgccttc
     4621 agtgctgagg cggcggtggc gcgcgccgcc aggcgggggc gaaggcactg tccgcggtgc
     4681 tgaagctggc agtgcgcacg cgcctcgccg catcctgttt cccctccccc tctctgatag
     4741 gggatgcgca atttggggaa tgggggttgg gtgcttgtcc agtgggtcgg ggtcggtcgt
     4801 caagcatgct ttaaatactg aaaagggcca attccagcac actggagctc ggatcttcct
     4861 agagcatggc tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct
     4921 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     4981 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     5041 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     5101 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     5161 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     5221 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     5281 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     5341 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     5401 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     5461 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     5521 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt
     5581 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     5641 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     5701 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     5761 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     5821 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     5881 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     5941 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     6001 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     6061 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     6121 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     6181 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     6241 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     6301 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     6361 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     6421 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     6481 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     6541 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     6601 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     6661 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     6721 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     6781 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     6841 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     6901 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     6961 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     7021 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     7081 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     7141 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     7201 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     7261 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     7321 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     7381 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     7441 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaag
//
