LOCUS       sequence-374363-        7028 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FLP-dependent expression of tdTomato under EF1a promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7028)
  AUTHORS   Xu J, Jo A, DeVries RP, Deniz S, Cherian S, Sunmola I, Song X, 
            Marshall JJ, Gruner KA, Daigle TL, Contractor A, Lerner TN, Zeng H, 
            Zhu Y
  TITLE     Intersectional mapping of multi-transmitter neurons and other cell 
            types in the brain.
  JOURNAL   Cell Rep. 2022 Jul 5;40(1):111036. doi: 
            10.1016/j.celrep.2022.111036.
  PUBMED    35793636
REFERENCE   2  (bases 1 to 7028)
  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: "Cell Rep.";
            date: "2022-07-5"; volume: "40(1):111036. doi"; pages: " 
            10.1016/j.celrep.2022.111036"
COMMENT     Sequence Label: pAAV-EF1a-fDIO-tdTomato-WPRE
FEATURES             Location/Qualifiers
     source          1..7028
                     /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    1440..1473
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(1570..3000)
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     regulatory      2997..3006
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3013..3046)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    3155..3743
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3208..3228)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3626..3637)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3775..4251
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3974..3993)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4291..4431
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4291..4420
                     /label=AAV2 ITR
     rep_origin      4506..4961
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4593..4612)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4803..4824
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4978..4997)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5097..5119
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5157..5175)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5243..5347
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5348..5416,5417..6208)
                     /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(5566..5585)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6379..6967
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6868..6887
                     /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 gactccggag
     1441 aagttcctat tctctagaaa gtataggaac ttcgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttagaagttc ctattcttca aaaggtatag gaacttcggc
     1561 gcgccattct tacttgtaca gctcgtccat gccgtacagg aacaggtggt ggcggccctc
     1621 ggagcgctcg tactgttcca cgatggtgta gtcctcgttg tgggaggtga tgtccagctt
     1681 ggtgtccacg tagtagtagc cgggcagttg cacgggcttc ttggccatgt agatggtctt
     1741 gaactccacc aggtagtggc cgccgtcctt cagcttcagg gcctggtgga tctcgccctt
     1801 cagcacgccg tcgcgggggt acaggcgctc ggtggaggcc tcccagccca tggtcttctt
     1861 ctgcattacg gggccgtcgg gggggaagtt ggtgccgcgc atcttcacct tgtagatcag
     1921 cgtgccgtcc tgcagggagg agtcctgggt cacggtcacc agaccgccgt cctcgaagtt
     1981 catcacgcgc tcccacttga agccctcggg gaaggacagc ttcttgtaat cggggatgtc
     2041 ggcggggtgc ttcacgtacg ccttggagcc gtacatgaac tggggggaca ggatgtccca
     2101 ggcgaagggc agggggccgc ccttggtcac cttcagcttg gcggtctggg tgccctcgta
     2161 ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcatgg agccctccat
     2221 gcgcaccttg aagcgcatga actctttgat gacggccatg ttgttgtcct cggaggaggc
     2281 ggtgccggag ctgccgctgc cggtgctgcc ggtgccatgc cccaggaaca ggtggtggcg
     2341 gccctcggag cgctcgtact gttccacgat ggtgtagtcc tcgttgtggg aggtgatgtc
     2401 cagcttggtg tccacgtagt agtagccggg cagttgcacg ggcttcttgg ccatgtagat
     2461 ggtcttgaac tccaccaggt agtggccgcc gtccttcagc ttcagggcct ggtggatctc
     2521 gcccttcagc acgccgtcgc gggggtacag gcgctcggtg gaggcctccc agcccatggt
     2581 cttcttctgc attacggggc cgtcgggggg gaagttggtg ccgcgcatct tcaccttgta
     2641 gatcagcgtg ccgtcctgca gggaggagtc ctgggtcacg gtcaccagac cgccgtcctc
     2701 gaagttcatc acgcgctccc acttgaagcc ctcggggaag gacagcttct tgtaatcggg
     2761 gatgtcggcg gggtgcttca cgtacgcctt ggagccgtac atgaactggg gggacaggat
     2821 gtcccaggcg aagggcaggg ggccgccctt ggtcaccttc agcttggcgg tctgggtgcc
     2881 ctcgtagggg cggccctcgc cctcgccctc gatctcgaac tcgtggccgt tcatggagcc
     2941 ctccatgcgc accttgaagc gcatgaactc tttgatgacc tcctcgccct tgctcaccat
     3001 ggtggcgcta gcgaagttcc tatactttct agagaatagg aacttcttgc cttaacccag
     3061 aaattatcac tgttattctt tagaatggtg caaagagaag ttcctatacc ttttgaagaa
     3121 taggaacttc gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg
     3181 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     3241 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     3301 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     3361 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     3421 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     3481 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     3541 gtcggggaaa tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg
     3601 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     3661 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     3721 ctccctttgg gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg
     3781 catccctgtg acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc
     3841 accagccttg tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat
     3901 aatattatgg ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt
     3961 agggcctgcg gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact
     4021 gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga
     4081 ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc
     4141 accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct
     4201 cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg
     4261 taggtaacca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc
     4321 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
     4381 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga
     4441 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
     4501 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     4561 gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct
     4621 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     4681 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
     4741 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     4801 tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga
     4861 tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa
     4921 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
     4981 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     5041 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     5101 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     5161 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     5221 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     5281 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     5341 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     5401 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     5461 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     5521 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     5581 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     5641 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     5701 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     5761 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     5821 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     5881 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     5941 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     6001 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     6061 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     6121 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     6181 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     6241 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     6301 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     6361 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     6421 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     6481 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     6541 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     6601 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6661 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     6721 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     6781 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     6841 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     6901 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     6961 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     7021 tcacatgt
//
