LOCUS       sequence-418114-        5314 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible AAV with reporter gene and cloning site to add activator 
            domains.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5314)
  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 5314)
  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: pJT360 AAV-Rapamycin Inducible-noActivator Dom...
FEATURES             Location/Qualifiers
     source          1..5314
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(162..181)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      975..1563
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1464..1483
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1625..1765
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1636..1765
                     /label=AAV2 ITR
     protein_bind    1772..1792
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     CDS             1947..1973
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2289..2342
                     /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             2343..2369
                     /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(2376..2549,2550..2555,2556..2738)
                     /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             2745..3068
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             2745..3065
                     /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        3270..3383
                     /label=minimal IL-2 promoter
                     /label=IL-2 promoter
                     /note="minimal promoter from the human interleukin-2 gene"
     regulatory      3387..3396
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3393..3395,3396..3398,3399..4109)
                     /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(3438..3459)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3699..3718)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4046..4067
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     repeat_region   4201..4341
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4201..4330
                     /label=AAV2 ITR
     rep_origin      4416..4871
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4503..4522)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4713..4734
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4888..4907)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5007..5029
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5067..5085)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5153..5257
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5258..5314,1..12,13..804)
                     /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 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
       61 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
      121 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
      181 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
      241 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
      301 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
      361 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
      421 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
      481 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
      541 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
      601 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtgga
      661 agccgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
      721 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
      781 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
      841 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
      901 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
      961 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     1021 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     1081 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     1141 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     1201 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     1261 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     1321 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     1381 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     1441 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     1501 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     1561 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     1621 atgtcctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg
     1681 cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg
     1741 gccaactcca tcactagggg ttccttctag acaactttgt atagaaaagt tgggtaccgc
     1801 ggccgcacgc gttaatgatg ggcgctacag taatgatggg cggaattcct cgaggtactt
     1861 atataagggg gtgggggcgc gttcgtcctc agtcgcgatc gaacactcga gccgagcaga
     1921 cgtgcctaac cggtgccgcc accatgcctg ctgccaagag ggtcaagttg gactggcatg
     1981 aaggcctgga agaggcatct cgtttgtact ttggggaaag gaacgtgaaa ggcatgtttg
     2041 aggtgctgga gcccttgcat gctatgatgg aacggggccc ccagactctg aaggaaacat
     2101 cttttaatca ggcctatggt cgtgatttaa tggaggccca agagtggtgc aggaagtaca
     2161 tgaaatcagg gaatgtcaag gacctcaccc aagcctggga cctctattat catgtgttcc
     2221 gacgaatctc aaagcagtct ggatccggat caggctccgg agacgggtca agcgtctctg
     2281 gatctggaga gggaagggga agcctcctaa cttgcggaga tgtcgaggag aatcctggcc
     2341 cacctgccgc taagcgtgtc aagttggact ctagagaacg cccgtatgct tgccctgtcg
     2401 agtcctgcga tcgccgcttt tctcgctcgg atgagcttac ccgccatatc cgcatccaca
     2461 caggccagaa gcccttccag tgtcgaatct gcatgcgtaa cttcagtcgt agtgaccacc
     2521 ttaccaccca catccgcacc cacacaggcg gcggccgcag gaggaagaaa cgcaccagca
     2581 tagagactaa catccgtgtg gccttagaga agagtttctt ggagaatcaa aagcctacct
     2641 cggaagagat cactatgatt gctgatcagc tcaatatgga aaaagaggtg attcgtgttt
     2701 ggttctgtaa ccgccgccag aaagaaaaaa gaatcaacaa gggcggcgtg caggtggaga
     2761 ctatctcccc aggcgacggg cgcaccttcc ccaagcgcgg ccagacctgc gtggtgcact
     2821 acaccgggat gcttgaagat ggaaagaaat ttgattcctc ccgggacaga aacaagccct
     2881 ttaagtttat gctaggcaag caggaggtga tccgaggctg ggaagaaggg gttgcccaga
     2941 tgagtgtggg tcagagagcc aaactgacta tatctccaga ttatgcctat ggtgccactg
     3001 ggcacccagg catcatccca ccacatgcca ctctcgtctt tgatgtggag cttctaaaac
     3061 tggaataaca tatgaaataa aatacgaaat gcgtacgtaa tgatgggcgc taaattaatg
     3121 atgggcggag ttgtaatgat gggcgctcgt ttaatgatgg gcgtctagct aatgatgggc
     3181 gggcgattaa tgatgggcga aggactaatg atgggcgcag cggtaatgat gggcgatcag
     3241 ctaatgatgg gcgacgcgaa ttaattcaac attttgacac ccccataata tttttccaga
     3301 attaacagta taaattgcat ctcttgttca agagttccct atcactctct ttaatcacta
     3361 ctcacagtaa cctcaactcc tgcgccgcca ccatggtgag caagggcgag gagctgttca
     3421 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     3481 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     3541 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccttcggc tacggcctga
     3601 tgtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     3661 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     3721 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     3781 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     3841 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     3901 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     3961 gcgacggccc cgtgctgctg cccgacaacc actacctgag ctaccagtcc gccctgagca
     4021 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     4081 tcactctcgg catggacgag ctgtacaagt aagcctaata aagagctcag atgcatgcat
     4141 cagagtgtgt tggttttttg tgtgtcggac cgagcggccg cgcatgctgg ggagggccgc
     4201 aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg
     4261 ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc
     4321 gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg
     4381 gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag
     4441 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc
     4501 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4561 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4621 aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     4681 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     4741 actcaactct atctcgggct attcttttga tttataaggg attttgccga tttcggtcta
     4801 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     4861 gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     4921 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     4981 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     5041 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     5101 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     5161 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     5221 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     5281 cgcccttatt cccttttttg cggcattttg cctt
//
