LOCUS       Exported                6200 bp ds-DNA     circular SYN 30-JUL-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6200)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Aug 2, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     UUID: 36b423e9-a453-4e2c-bbd4-57ba04c575d9
            Type: Synthetic
            ConfirmedExperimentally: 0
            CustomMapLabel: pAAV-hsyn-GRAB_DA2m
            UseCustomMapLabel: 1
            Description: Expresses the 2nd generation medium-affinity DA sensor 
            GRAB_DA2m in neurons.
            SequenceClass: UNA
            TransformedInto: unspecified
            References: Reference: type=""Journal Article"" 
            title=""Next-generation GRAB sensors for monitoring dopaminergic 
            activity in vivo."" journalName=""Nature Methods (2020)"" 
            journal=""Nature Methods (2020)"" authors=""Sun F, Zhou J, Dai B, 
            Qian T, Zeng J, Li X, Zhuo Y, Zhang Y, Wang Y, Qian C, Tan K, Feng 
            J, Dong H, Lin D, Cui G, Li Y.""
FEATURES             Location/Qualifiers
     source          1..6200
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /locus_tag="AAV2 ITR (alternate)"
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      581..586
                     /locus_tag="Kozak sequence"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
     misc_feature    590..2299
                     /locus_tag="iPASnFR-L"
                     /label=iPASnFR-L
     misc_feature    2327..2915
                     /locus_tag="WPRE"
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     primer_bind     2380..2400
                     /locus_tag="WPRE-R"
                     /PCR_conditions="primer sequence:CATAGCGTAAAAGGAGCAACA"
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             2798..2809
                     /codon_start=1
                     /locus_tag="Factor Xa site"
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="SALN"
     polyA_signal    2947..3423
                     /locus_tag="hGH poly(A) signal"
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     3146..3165
                     /locus_tag="hGH-PA-R"
                     /PCR_conditions="primer sequence:CCAGCTTGGTTCCCAATAGA"
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3463..3603
                     /locus_tag="AAV2 ITR(1)"
                     /label=AAV2 ITR
                     /label=AAV2 ITR(1)
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3463..3592
                     /locus_tag="AAV2 ITR"
                     /label=AAV2 ITR
     rep_origin      3678..4133
                     /direction=RIGHT
                     /locus_tag="f1 ori"
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     3765..3784
                     /locus_tag="F1ori-R"
                     /PCR_conditions="primer sequence:AGGGAAGAAAGCGAAAGGAG"
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3975..3996
                     /locus_tag="F1ori-F"
                     /PCR_conditions="primer sequence:GTGGACTCTTGTTCCAAACTGG"
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4150..4169
                     /locus_tag="pRS-marker"
                     /PCR_conditions="primer sequence:CGGCATCAGAGCAGATTGTA"
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable
                     marker"
     primer_bind     4269..4291
                     /locus_tag="pGEX 3'"
                     /PCR_conditions="primer sequence:CCGGGAGCTGCATGTGTCAGAGG"
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4329..4347
                     /locus_tag="pBRforEco"
                     /PCR_conditions="primer sequence:AATAGGCGTATCACGAGGC"
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward
                     primer"
     promoter        4415..4519
                     /gene="bla"
                     /locus_tag="AmpR promoter"
                     /label=AmpR promoter
     CDS             4520..5380
                     /codon_start=1
                     /gene="bla"
                     /locus_tag="AmpR"
                     /product="I²-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin,
                     and related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4738..4757
                     /locus_tag="Amp-R"
                     /PCR_conditions="primer sequence:ATAATACCGCGCCACATAGC"
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5551..6139
                     /direction=RIGHT
                     /locus_tag="ori"
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5892..5901
                     /locus_tag="F1ori-F(1)"
                     /PCR_conditions="primer sequence:GTGGACTCTTGTTCCAAACTGG"
                     /label=F1ori-F
                     /label=F1ori-F(1)
                     /note="F1 origin, forward primer"
     primer_bind     6040..6059
                     /locus_tag="pBR322ori-F"
                     /PCR_conditions="primer sequence:GGGAAACGCCTGGTATCTTT"
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtatgagg cggggtgggg gtgcctacct gacgaccgac
      181 cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag
      241 agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg
      301 acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg
      361 ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc
      421 gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc
      481 gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga
      541 ggagtcgtgt cgtgcctgag agcgcagtcg agaaaccggt gccaccatga acacgttgac
      601 ggtttacaat tggggtgatt acatcgaccc tgcattaatc cgtaagttcg agaagcaaac
      661 gggtataaag gttatatatc agacgtttga ctccaatgaa gcaatgatgg caaagattgc
      721 tcaaggtgga gcaacgttcg acgtagcagt tccatctgag tacgcgatct ctaagatgaa
      781 ggaagatgga ttgctgatcc ccttagacca ttcaaaatta cctaacctca aatacattaa
      841 tccgcgcttc ttaaatttaa gcttcgaccc cggaaacaga tactctgtcc cctacttctg
      901 gggtaccgtg gggatagttt acaatcccgg catgttgggt ggtaagaaga tcacttcttg
      961 gaacgacttg tgggatccag acctgcgcaa tcagatcctg cttgttgacg gagctcgtgc
     1021 ggtaatcgga atgggactta actcactcca ctattcactc aacgacacca ataaagagca
     1081 tctcaagaag gccaaagaga agttggacag actcgtgcca aatgtgaaag ccatagtcgg
     1141 ggatgaaatc aagatgttgc tggcgaatga ggaagcagcg ataggagttg tgtggagtgg
     1201 ggacgctgcg gagatcatcg ccgaaaatga caagttggat tacgtggtgc ccaaagaggg
     1261 atcgaacttg tggttcgaca atatggtgat tccaaagacc gcgaagaata ttgaaggagc
     1321 acataaattc attaacttca tgctggaccc aaagaatgcg gcgcagaacg ccgagtttgt
     1381 tggctattca accccaaata agaaggcttt ggattacatg cctaaggaaa tcacagagtc
     1441 taagaggcac acggtctata tcaccgccga caagcagaag aacggcatca aggcgaactt
     1501 caagatccac cttaacgtgg aggacggcag cgtgcagctc gccgaccact accagcagaa
     1561 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gctatcagtc
     1621 cgtgctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     1681 cgccaccggg aactctctcg gcatggacga gctgtacaag ggcggtaccg gagggagcat
     1741 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1801 cgtaaacggc cacaagttca gcgtgcgcgg cgagggcgag ggcgatgcca ccaacggcaa
     1861 gctaaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1921 gaccaccctg agctacggcg tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
     1981 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcagcttcaa
     2041 ggacgacgga acctacaaga cccgcgccga ggtgaagtta gagggcgaca ccctggtgaa
     2101 ccgcatcgag ctaaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     2161 ggagtacaac tttaactatc atgacaagcg gttctatccc gatttcgact caatcggcaa
     2221 tttagaagtt tacaagaatc ttggaaagcg tatgttggct tactacaatg aactgttcct
     2281 gcaatttaag atgcaccgct aagaattcga tatcaagctt atcgataatc aacctctgga
     2341 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg
     2401 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt
     2461 ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag
     2521 gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc
     2581 caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga
     2641 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
     2701 ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac
     2761 ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct
     2821 tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca
     2881 gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag
     2941 atctacgggt ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc
     3001 actccagtgc ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag
     3061 gtgtccttct ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg
     3121 aagacaacct gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa
     3181 tcttggctca ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc
     3241 gagttgttgg gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag
     3301 agacggggtt tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac
     3361 ccaccttggc ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc
     3421 cttctgattt tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg
     3481 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     3541 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc
     3601 aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     3661 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     3721 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt
     3781 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     3841 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
     3901 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     3961 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg
     4021 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct
     4081 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
     4141 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     4201 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     4261 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     4321 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     4381 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     4441 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     4501 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     4561 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     4621 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     4681 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     4741 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     4801 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     4861 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     4921 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     4981 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5041 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     5101 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     5161 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     5221 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     5281 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     5341 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     5401 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     5461 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     5521 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     5581 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     5641 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     5701 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     5761 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     5821 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     5881 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     5941 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     6001 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     6061 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     6121 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     6181 ctggcctttt gctcacatgt
//
