LOCUS       sequence-418115-        5512 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible AAV activator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5512)
  AUTHORS   Tycko J, Van MV, Aradhana, DelRosso N, Ye H, Yao D, Valbuena R, 
            Vaughan-Jackson A, Xu X, Ludwig C, Spees K, Liu K, Gu M, Khare V, 
            Mukund AX, Suzuki PH, Arana S, Zhang C, Du PP, Ornstein TS, Hess GT,
            Kamber RA, Qi LS, Khalil AS, Bintu L, Bassik MC
  TITLE     Development of compact transcriptional effectors using 
            high-throughput measurements in diverse contexts.
  JOURNAL   Nat Biotechnol. 2024 Nov 1. doi: 10.1038/s41587-024-02442-6.
  PUBMED    39487265
REFERENCE   2  (bases 1 to 5512)
  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: "Nat 
            Biotechnol. 2024 Nov 1. doi: 10.1038/s41587-024-02442-6."
COMMENT     Sequence Label: pJT314 AAV-Rapamycin Inducible-NZ-Citrine
FEATURES             Location/Qualifiers
     source          1..5512
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(121..140)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      934..1522
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1423..1442
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1584..1713
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             1862..1888
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2477..2530
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2531..2557
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             join(2564..2737,2738..2743,2744..2926)
                     /codon_start=1
                     /product="composite human DNA-binding domain composed of 
                     domains from the transcription factors Zif268 and Oct-1 
                     (Pomerantz et al., 1995)"
                     /label=ZFHD1
                     /label=ZFHD1 (DNA binding domain)
                     /translation="ERPYACPVESCDRRFSRSDELTRHIRIHTGQKPFQCRICMRNFSR
                     SDHLTTHIRTHTGGGRRRKKRTSIETNIRVALEKSFLENQKPTSEEITMIADQLNMEKE
                     VIRVWFCNRRQKEKRIN"
     CDS             2933..3256
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             2933..3253
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=DmrB
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     promoter        3458..3571
                     /label=minimal IL-2 promoter
                     /label=IL-2 promoter
                     /note="minimal promoter from the human interleukin-2 gene"
     regulatory      3575..3584
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3581..3583,3584..3586,3587..4297)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3626..3647)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3887..3906)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4234..4255
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     repeat_region   4370..4510
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4370..4499
                     /label=AAV2 ITR
     rep_origin      4585..5040
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4672..4691)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4882..4903
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5057..5076)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5170..5192
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5224..5242)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5310..5414
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5415..5483,5484..5512,1..763)
                     /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 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
       61 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
      121 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
      181 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
      241 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
      301 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
      361 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
      421 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
      481 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
      541 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
      601 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
      661 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
      721 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
      781 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
      841 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
      901 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
      961 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     1021 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     1081 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     1141 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     1201 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     1261 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     1321 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     1381 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     1441 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     1501 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     1561 ttgctggcct tttgctcaca tgtcctgcag gcagctgcgc gctcgctcgc tcactgaggc
     1621 cgcccgggcg tcgggcgacc tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag
     1681 agggagtggc caactccatc actaggggtt cctgcggccg cacgcgttaa tgatgggcgc
     1741 tacagtaatg atgggcggaa ttcctcgagg tacttatata agggggtggg ggcgcgttcg
     1801 tcctcagtcg cgatcgaaca ctcgagccga gcagacgtgc ctaaccggtg ccgccaccat
     1861 gcctgctgcc aagagggtca agttggactg gcatgaaggc ctggaagagg catctcgttt
     1921 gtactttggg gaaaggaacg tgaaaggcat gtttgaggtg ctggagccct tgcatgctat
     1981 gatggaacgg ggcccccaga ctctgaagga aacatctttt aatcaggcct atggtcgtga
     2041 tttaatggag gcccaagagt ggtgcaggaa gtacatgaaa tcagggaatg tcaaggacct
     2101 cacccaagcc tgggacctct attatcatgt gttccgacga atctcaaagc agtctggatc
     2161 cggatcaggc tccgaaggcc aatcagatga aagggcgttg ctcgatcaac ttcatactct
     2221 cctttccaat acagatgcta caggtctcga agaaattgat cgggcactcg gaattcctga
     2281 acttgttaat caaggtcaag ctctcgaacc aaaacaaggt ggggggggaa gtggctttgt
     2341 aactctcaaa gatgttggta tggattttac tcttggggat tgggaacaat tggggctcga
     2401 acaaggagat acattttggg atacagctct tgataattgt caagatctct tcttgctcga
     2461 tcccccagga tctggagagg gaaggggaag cctcctaact tgcggagatg tcgaggagaa
     2521 tcctggccca cctgccgcta agcgtgtcaa gttggactct agagaacgcc cgtatgcttg
     2581 ccctgtcgag tcctgcgatc gccgcttttc tcgctcggat gagcttaccc gccatatccg
     2641 catccacaca ggccagaagc ccttccagtg tcgaatctgc atgcgtaact tcagtcgtag
     2701 tgaccacctt accacccaca tccgcaccca cacaggcggc ggccgcagga ggaagaaacg
     2761 caccagcata gagactaaca tccgtgtggc cttagagaag agtttcttgg agaatcaaaa
     2821 gcctacctcg gaagagatca ctatgattgc tgatcagctc aatatggaaa aagaggtgat
     2881 tcgtgtttgg ttctgtaacc gccgccagaa agaaaaaaga atcaacaagg gcggcgtgca
     2941 ggtggagact atctccccag gcgacgggcg caccttcccc aagcgcggcc agacctgcgt
     3001 ggtgcactac accgggatgc ttgaagatgg aaagaaattt gattcctccc gggacagaaa
     3061 caagcccttt aagtttatgc taggcaagca ggaggtgatc cgaggctggg aagaaggggt
     3121 tgcccagatg agtgtgggtc agagagccaa actgactata tctccagatt atgcctatgg
     3181 tgccactggg cacccaggca tcatcccacc acatgccact ctcgtctttg atgtggagct
     3241 tctaaaactg gaataacata tgaaataaaa tacgaaatgc gtacgtaatg atgggcgcta
     3301 aattaatgat gggcggagtt gtaatgatgg gcgctcgttt aatgatgggc gtctagctaa
     3361 tgatgggcgg gcgattaatg atgggcgaag gactaatgat gggcgcagcg gtaatgatgg
     3421 gcgatcagct aatgatgggc gacgcgaatt aattcaacat tttgacaccc ccataatatt
     3481 tttccagaat taacagtata aattgcatct cttgttcaag agttccctat cactctcttt
     3541 aatcactact cacagtaacc tcaactcctg cgccgccacc atggtgagca agggcgagga
     3601 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa
     3661 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt
     3721 catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccttcggcta
     3781 cggcctgatg tgcttcgccc gctaccccga ccacatgaag cagcacgact tcttcaagtc
     3841 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta
     3901 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa
     3961 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa
     4021 cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa
     4081 gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac
     4141 ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagct accagtccgc
     4201 cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc
     4261 cgccgggatc actctcggca tggacgagct gtacaagtaa gcctaataaa gagctcagat
     4321 gcatgcatca gagtgtgttg gttttttgtg tgtcggaccg agcggccgca ggaaccccta
     4381 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
     4441 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
     4501 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     4561 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     4621 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg
     4681 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     4741 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     4801 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     4861 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     4921 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     4981 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     5041 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     5101 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     5161 aagctgtgac cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcaa
     5221 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     5281 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5341 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5401 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5461 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa ga
//
