LOCUS       sequence-487698-        6720 bp    DNA     circular SYN 02-OCT-2025
DEFINITION  Express Kir 2.1 and dTomato in the neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6720)
  AUTHORS   Li H, Su C, Xu Y, Ludwig MQ, Davis J, Tong Q
  TITLE     An alternative neural basis underlying leptin resistance.
  JOURNAL   Cell Rep. 2025 Jun 19;44(7):115863. doi: 
            10.1016/j.celrep.2025.115863.
  PUBMED    40540394
REFERENCE   2  (bases 1 to 6720)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 2, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Rep.";
            date: "2025-06-19"; volume: "44(7):115863. doi"; pages: " 
            10.1016/j.celrep.2025.115863"
COMMENT     Sequence Label: pAAV-SYN1>Fas-Myc/Kir2.1*(E224G Y242F):P2A :dT...
FEATURES             Location/Qualifiers
     source          1..6720
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        166..613
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      686..695
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             695..724
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2021..2077
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2078..2779
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(2211..2229)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2479..2498
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2673..2696
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     misc_feature    2847..3435
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2900..2920)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3318..3329)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3467..3943
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3666..3685)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3983..4123
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3983..4112
                     /label=AAV2 ITR
     rep_origin      4198..4653
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4285..4304)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4495..4516
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4670..4689)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4789..4811
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4849..4867)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4935..5039
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5040..5108,5109..5900)
                     /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(5258..5277)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6071..6659
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6560..6579
                     /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 gcgtctgcag agggccctgc gtatgagtgc aagtgggttt
      181 taggaccagg atgaggcggg gtgggggtgc ctacctgacg accgaccccg acccactgga
      241 caagcaccca acccccattc cccaaattgc gcatccccta tcagagaggg ggaggggaaa
      301 caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca ccgcggacag tgccttcgcc
      361 cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa ggcgcgctga cgtcactcgc
      421 cggtcccccg caaactcccc ttcccggcca ccttggtcgc gtccgcgccg ccgccggccc
      481 agccggaccg caccacgcga ggcgcgagat aggggggcac gggcgcgacc atctgcgctg
      541 cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc agcggaggag tcgtgtcgtg
      601 cctgagagcg cagggtaccg gatcctctag agtcgactcc ggaacaactt cgtatatacc
      661 tttctatacg aagttgtggc gcgccgccac catggagcag aagctgatca gcgaggagga
      721 cctgggcagt gtgagaacca accggtacag cattgtctct tcggaggaag atggcatgaa
      781 gctggccact atggcagttg ccaatggctt tgggaatggc aagagtaaag tccatacccg
      841 acaacagtgc aggagccgct ttgtgaagaa agatggtcat tgcaatgttc agtttatcaa
      901 cgtgggtgag aagggacaga ggtacctggc agacatcttt actacctgtg tcgacatccg
      961 ctggaggtgg atgctggtta tcttctgtct tgccttcgtg ctctcctggc tgttctttgg
     1021 ctgtgtgttt tggttgatag ccctgctcca tggggatcta gatacttcta aagtgagcaa
     1081 agcatgcgtg tcggaggtta acagcttcac ggctgccttc ctcttctcca tcgagaccca
     1141 gacaaccatt ggctatggtt tcaggtgtgt gacagacgaa tgcccaattg ctgtcttcat
     1201 ggtggtattc cagtcaattg taggctgcat cattgacgcc ttcatcattg gtgcagtcat
     1261 ggcgaagatg gcaaagccaa agaagagaaa tgagactctt gtcttcagtc acaatgctgt
     1321 gattgccatg agggatggca aactctgctt gatgtggaga gtgggtaacc ttcgaaagag
     1381 ccaccttgtg ggagctcatg tccgggcaca gcttctcaaa tctaggatca cttcagaagg
     1441 ggagtttatc cctttggacc agatagacat caatgttggt tttgatagtg gaattgaccg
     1501 catatttcta gtgtccccca tcactatcgt tcacgaaata gatgaagaca gccctttata
     1561 tgacttgagt aagcaggaca ttgacaatgc agactttgaa attgttgtca tactggaagg
     1621 catggtggag gcgactgcca tgacaactca atgccggagt tcgtatctgg ccaatgaaat
     1681 tctctggggt caccgctatg agccagtgct ctttgaagag aaacactact ataaagtaga
     1741 ctattcaaga ttccataaga cttatgaagt acctaacacc cccctttgta gtgccagaga
     1801 cttagcagag aagaaataca tcctttcaaa tgcaaattca ttttgctatg aaaatgaagt
     1861 tgccctaaca agcaaagagg aagaggagga tagtgagaac ggagtcccag agagcacaag
     1921 cacagactca cctcctggca tagatctcca caaccaggca agcgtacctc tagagcccag
     1981 gcccttaagg cgagaatcgg agatagataa cggaagcgga gccactaact tctccctgtt
     2041 gaaacaagca ggggatgtcg aagagaatcc cgggccaatg gtgagcaagg gcgaggaggt
     2101 catcaaagag ttcatgcgct tcaaggtgcg catggagggc tccatgaacg gccacgagtt
     2161 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa
     2221 ggtgaccaag ggcggccccc tgcccttcgc ctgggacatc ctgtcccccc agttcatgta
     2281 cggctccaag gcgtacgtga agcaccccgc cgacatcccc gattacaaga agctgtcctt
     2341 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggtc tggtgaccgt
     2401 gacccaggac tcctccctgc aggacggcac gctgatctac aaggtgaaga tgcgcggcac
     2461 caacttcccc cccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctccac
     2521 cgagcgcctg tacccccgcg acggcgtgct gaagggcgag atccaccagg ccctgaagct
     2581 gaaggacggc ggccactacc tggtggagtt caagaccatc tacatggcca agaagcccgt
     2641 gcaactgccc ggctactact acgtggacac caagctggac atcacctccc acaacgagga
     2701 ctacaccatc gtggaacagt acgagcgctc cgagggccgc caccacctgt tcctgtacgg
     2761 catggacgag ctgtacaagt aagctagcac aacttcgtat atacctttct atacgaagtt
     2821 gtgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
     2881 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     2941 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     3001 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     3061 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     3121 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     3181 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     3241 aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg cgcgggacgt
     3301 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     3361 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     3421 gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt ggcatccctg
     3481 tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc ccaccagcct
     3541 tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct ataatattat
     3601 ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct gtagggcctg
     3661 cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca ctgcaatctc
     3721 cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg gattccaggc
     3781 atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt tcaccatatt
     3841 ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc ctcccaaatt
     3901 gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt tgtaggtaac
     3961 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     4021 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4081 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4141 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4201 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4261 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4321 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4381 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     4441 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4501 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     4561 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4621 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     4681 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     4741 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     4801 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     4861 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     4921 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     4981 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5041 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5101 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5161 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5221 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5281 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5341 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5401 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     5461 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     5521 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     5581 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     5641 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     5701 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     5761 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     5821 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     5881 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     5941 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6001 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6061 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6121 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6181 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     6241 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6301 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6361 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     6421 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     6481 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     6541 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     6601 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     6661 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
