LOCUS       sequence-432134-        5591 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Provides AND intersectional (Cre+Flp) expression of TVA-mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5591)
  AUTHORS   Hughes AC, Pollard BG, Xu B, Gammons JW, Chapman P, Bikoff JB, 
            Schwarz LA
  TITLE     A Novel Single Vector Intersectional AAV Strategy for Interrogating 
            Cellular Diversity and Brain Function.
  JOURNAL   bioRxiv. 2023 Feb 8:2023.02.07.527312. doi: 
            10.1101/2023.02.07.527312. Preprint.
  PUBMED    36798174
REFERENCE   2  (bases 1 to 5591)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv. 
            2023 Feb 8:2023.02.07.527312. doi: 10.1101/2023.02.07.527312. 
            Preprint."
COMMENT     Sequence Label: pAAV-hSyn-ConVERGD-TVA(mChr)-W3SL
FEATURES             Location/Qualifiers
     source          1..5591
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(101..120)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      914..1502
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1403..1422
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    1790..1811
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1826..1831,1832..1849,1850..1856)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1864..1880
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     1869..1891
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1888..1904
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1888..1904
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   1944..2084
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1955..2084
                     /label=AAV2 ITR
     primer_bind     2457..2477
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    2581..2628
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     regulatory      2743..2752
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3187..3894
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(3329..3347)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3597..3616
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    3915..3948
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4041..4074
                     /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(4087..4134)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        complement(4243..4690)
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     repeat_region   4724..4853
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     primer_bind     complement(4879..4896)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4879..4895)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4888..4910)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5104..5123)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5223..5245
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5283..5301)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5369..5473
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5474..5542,5543..5591,1..743)
                     /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"
ORIGIN
        1 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
       61 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
      121 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
      181 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
      241 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
      301 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
      361 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
      421 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
      481 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
      541 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtggtt
      601 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
      661 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
      721 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
      781 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
      841 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
      901 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
      961 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     1021 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     1081 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     1141 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     1201 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     1261 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     1321 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     1381 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     1441 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     1501 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     1561 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     1621 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     1681 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
     1741 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
     1801 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
     1861 gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc
     1921 tcggcgcgcc attgggatcc aatcctgcag gcagctgcgc gctcgctcgc tcactgaggc
     1981 cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg
     2041 agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggagct ccggttaccg
     2101 gccgctttaa aaaacctccc acatctcccc ctgaacctga aacataaaat gaatgcaatt
     2161 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     2221 aatttcacaa ataaagctag atggttacaa ataaagcaat agcatcacaa atttcacaaa
     2281 taaacaccac ggaattgtca gtgcccaaca gccgagcccc tgtccagcag cgggcaaggc
     2341 aggcggcgat gagttccgcc gtggcaagaa ctaaccagga tttatacaag gaggagaaaa
     2401 tgaaagccat acgggaagca atagcatgat acaaaggcat taaagcagcg tatccacata
     2461 gcgtaaaagg agcaacatag ttaagaatac cagtcaatct ttcacaaatt ttgtaatcca
     2521 gaggttgatt atctcgggca tcgaatatcg aatcggctaa gctctagaaa gcttcgtacg
     2581 gaagttacta ttccgaagtt cctattctct agaaagtata ggaacttcgt gtcgatggtg
     2641 atgcttggca attcgggtcg atggtgaagc attggtcttc ctattttgat gaagttccta
     2701 ttcttcaaaa ggtataggaa cttcctcgag accggtacta gtgccaccat ggcgcggctg
     2761 ctgcccgcgc tgctgctgct gctgctgccc ggtaacgtga ccggtaacgg gtccggtaac
     2821 ggttctttgt cccgttgccc ccccggtcag ttccgctgct cggagccgcc cggtgcccac
     2881 ggggagtgtt acccgcagga ctggctgtgc gacggacacc ccgactgcga cgacgggcgg
     2941 gacgagtggg gctgcgggac cagcgcgatc cccgcggtgc ccacggacaa cggcacagag
     3001 gctcccactg cccctgctcc tggacgtgct ctgccagcca ggaatcacgg ccgcatgtgg
     3061 atgctgatca ctgcagtgct cctgtgctgc ctggtagctg tgggtggtat cgctgcatgg
     3121 gggaagtcca aagcaaaaag caggtctgac atcttcagtc ttgaaagcgc atccaagggg
     3181 ggtggggtga gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc
     3241 aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag
     3301 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg
     3361 cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag
     3421 caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag
     3481 cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag
     3541 gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc
     3601 gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac
     3661 ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac
     3721 gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac
     3781 gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac
     3841 gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa gtaagcatgc
     3901 acatgcatga attcataact tcgtataatg tatgctatac gaagttattt tctttctttg
     3961 cgcgtcctgg attcgcggaa acgcgtacat ccagctgacg agtcccaaat aggacgaaac
     4021 gcgcttttct ttttgttttt ataacttcgt ataatgtatg ctatacgaag ttatcttaag
     4081 gtcgacgaag ttcctatact ttctagagaa taggaacttc ggaataggaa cttcgtgtcg
     4141 atggtgatgc ttggcaattc gggtcgatgg tgaagcattg gtcttcctat tttgatgaag
     4201 ttcctatacc ttttgaagaa taggaacttc gctagcggta ccctgcgctc tcaggcacga
     4261 cacgactcct ccgctgccca ccgcagactg aggcagcgct gagtcgccgg cgccgcagcg
     4321 cagatggtcg cgcccgtgcc cccctatctc gcgcctcgcg tggtgcggtc cggctgggcc
     4381 ggcggcggcg cggacgcgac caaggtggcc gggaagggga gtttgcgggg gaccggcgag
     4441 tgacgtcagc gcgccttcag tgctgaggcg gcggtggcgc gcgccgccag gcgggggcga
     4501 aggcactgtc cgcggtgctg aagctggcag tgcgcacgcg cctcgccgca tcctgtttcc
     4561 cctccccctc tctgataggg gatgcgcaat ttggggaatg ggggttgggt gcttgtccag
     4621 tgggtcgggg tcggtcgtca ggtaggcacc cccaccccgc ctcatcctgg tcctaaaacc
     4681 cacttgcact gcgatcgcgg gcccgtgcag catctgcggc cgcaggaacc cctagtgatg
     4741 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     4801 gcccgacgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggattggat
     4861 cccaatacgc gtcaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     4921 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     4981 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct
     5041 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct
     5101 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     5161 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgacagt
     5221 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacaaaa
     5281 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     5341 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     5401 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5461 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5521 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5581 tcagttgggt g
//
