LOCUS       sequence-190086-        7148 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependently express GFPL10 fusion using Introvert Switch.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7148)
  AUTHORS   Nectow AR, Schneeberger M, Zhang H, Field BC, Renier N, Azevedo E, 
            Patel B, Liang Y, Mitra S, Tessier-Lavigne M, Han MH, Friedman JM
  TITLE     Identification of a Brainstem Circuit Controlling Feeding.
  JOURNAL   Cell. 2017 Jul 27;170(3):429-442.e11. doi: 
            10.1016/j.cell.2017.06.045.
  PUBMED    28753423
REFERENCE   2  (bases 1 to 7148)
  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: 
            "10.1016/j.cell.2017.06"; journalName: "Cell"; date: "2017-07-27- 
            27"; volume: "170"; issue: "3"; pages: "429-442.e11"
COMMENT     Sequence Label: pAAV-IV-GFPL10
FEATURES             Location/Qualifiers
     source          1..7148
                     /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        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1440..1473)
                     /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(1524..1557)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(1617..2327,2328..2330,2331..2333))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1617..2327,2328..2330,2331..2333))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1659..1680)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2008..2027
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2267..2288
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2330..2339
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2344..2377
                     /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    2428..2461
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3275..3863
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3328..3348)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3746..3757)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3895..4371
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4094..4113)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4411..4551
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4411..4540
                     /label=AAV2 ITR
     rep_origin      4626..5081
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4713..4732)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4923..4944
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5098..5117)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5217..5239
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5277..5295)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5363..5467
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5468..5536,5537..6328)
                     /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(5686..5705)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6499..7087
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6988..7007
                     /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 gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa
     1441 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
     1561 gcgcctacgc ggccgccctg tcttgtaacc ttgatactta cctgagtccg gccggacttg
     1621 tacagctcgt ccatgccgag agtgatcccg gcggcggtca cgaactccag caggaccatg
     1681 tgatcgcgct tctcgttggg gtctttgctc agggcggact gggtgctcag gtagtggttg
     1741 tcgggcagca gcacggggcc gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc
     1801 tgcacgctgc cgtcctcgat gttgtggcgg atcttgaagt tcaccttgat gccgttcttc
     1861 tgcttgtcgg ccatgatata gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc
     1921 aggatgttgc cgtcctcctt gaagtcgatg cccttcagct cgatgcggtt caccagggtg
     1981 tcgccctcga acttcacctc ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg
     2041 gtgcgctcct ggacgtagcc ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg
     2101 tggtcggggt agcggctgaa gcactgcacg ccgtaggtca gggtggtcac gagggtgggc
     2161 cagggcacgg gcagcttgcc ggtggtgcag atgaacttca gggtcagctt gccgtaggtg
     2221 gcatcgccct cgccctcgcc ggacacgctg aacttgtggc cgtttacgtc gccgtccagc
     2281 tcgaccagga tgggcaccac cccggtgaac agctcctcgc ccttgctcac catggtggct
     2341 agcataactt cgtataaagt atcctatacg aagttatttg ccttaaccca gaaattatca
     2401 ctgttattct ttagaatggt gcaaagaata acttcgtata atgtatgcta tacgaagtta
     2461 tcaattgttt aaggagacca atagaaactg ggcttgtcga gacagagaag actcttgcgt
     2521 ttctgatagg cacctattgg tcttactgac atccactttg cctttctctc cacagatctc
     2581 gagctcaagc ttcgaattta gcagcaaagt ctcacgcgac accctgtacg aggcggtgcg
     2641 ggaagtcctg cacgggaacc agcgcaagcg ccgcaagttt ctggagacgg tggagctgca
     2701 gatcagcctg aagaactacg accctcagaa ggacaaacgt ttctcgggca ccgtcaggct
     2761 caagtccacc ccacgcccca agttctcggt gtgcgttctg ggggaccagc agcactgtga
     2821 tgaagccaag gccgtggata tcccccacat ggacatcgag gcgctcaaga agcttaacaa
     2881 aaacaagaag ttggtcaaga agctggctaa gaagtacgat gcctttttgg cctctgagtc
     2941 tctgattaag cagatcccac gtatcctggg cccaggccta aacaaggctg gcaagttccc
     3001 ctccctgctg acacacaatg aaaacatggt ggccaaagtg gatgaggtga aatcgacaat
     3061 caagttccag atgaagaagg tgctgtgttt ggccgtcgct gttggccacg tgaagatgac
     3121 cgatgatgag ctagtctaca acattcatct ggctgtcaat ttcttggtgt ccttgcttaa
     3181 gaaaaactgg caaaacgtgc gggctctgta catcaagagc accatgggca agccccagcg
     3241 tctgtattag gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg
     3301 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     3361 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     3421 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     3481 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     3541 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     3601 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     3661 gtcggggaaa tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg
     3721 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     3781 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     3841 ctccctttgg gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg
     3901 catccctgtg acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc
     3961 accagccttg tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat
     4021 aatattatgg ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt
     4081 agggcctgcg gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact
     4141 gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga
     4201 ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc
     4261 accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct
     4321 cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg
     4381 taggtaacca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc
     4441 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
     4501 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga
     4561 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
     4621 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     4681 gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct
     4741 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     4801 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
     4861 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     4921 tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga
     4981 tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa
     5041 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
     5101 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     5161 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     5221 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     5281 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     5341 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     5401 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     5461 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     5521 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     5581 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     5641 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     5701 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     5761 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     5821 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     5881 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     5941 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     6001 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     6061 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     6121 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     6181 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     6241 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     6301 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     6361 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     6421 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     6481 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     6541 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     6601 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     6661 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     6721 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6781 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     6841 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     6901 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     6961 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     7021 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     7081 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     7141 tcacatgt
//
