LOCUS       sequence-429969-        7616 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tet-inducible Cal-ID (V5) for Sleeping Beauty transposon system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7616)
  AUTHORS   Kim JW, Yong AJH, Aisenberg EE, Lobel JH, Wang W, Dawson TM, Dawson 
            VL, Gao R, Jan YN, Bateup HS, Ingolia NT
  TITLE     Molecular recording of calcium signals via calcium-dependent 
            proximity labeling.
  JOURNAL   Nat Chem Biol. 2024 Jul;20(7):894-905. doi: 
            10.1038/s41589-024-01603-7. Epub 2024 Apr 24.
  PUBMED    38658655
REFERENCE   2  (bases 1 to 7616)
  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: 
            "10.1038/s41589-024-01603-7"; journalName: "Nat Chem Biol"; date: 
            "2024-07"; volume: "20"; issue: "7"; pages: "894-905"
COMMENT     Sequence Label: pNTI821
FEATURES             Location/Qualifiers
     source          1..7616
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        295..607
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    303..321
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    339..357
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    374..392
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    410..428
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    446..464
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    481..499
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    517..535
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     580..604
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             731..787
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     CDS             join(788..796,797..811)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             2246..2287
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     polyA_signal    2326..2447
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2337..2356)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2391..2410
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             join(3170..3787,3788..3793,3794..3829,3830..3832,3833..3868,
                     3869..3871,3872..3907,3908..3913)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     CDS             3920..3976
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             3977..5002
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3977..5002
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     primer_bind     complement(5057..5074)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5063..5287
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5613..5630)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5784..6372)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5864..5883)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6543..7334,7335..7403))
                     /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"
     promoter        complement(7404..7508)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggatccctat acagttgaag tcggaagttt acatacactt aagttggagt cattaaaact
       61 cgtttttcaa ctactccaca aatttcttgt taacaaacaa tagttttggc aagtcagtta
      121 ggacatctac tttgtgcatg acacaagtca tttttccaac aattgtttac agacagatta
      181 tttcacttat aattcactgt atcacaattc cagtgggtca gaagtttaca tacactaagt
      241 tcgactcctc tgcagaatgc ggcgatgttt cggtaagggg tccgctatct agacgagttt
      301 actccctatc agtgatagag aacgtatgtc gagtttactc cctatcagtg atagagaacg
      361 atgtcgagtt tactccctat cagtgataga gaacgtatgt cgagtttact ccctatcagt
      421 gatagagaac gtatgtcgag tttactccct atcagtgata gagaacgtat gtcgagttta
      481 tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgta
      541 tgtcgattag gcgtgtacgg tgggcgccta taaaagcaga gctcgtttag tgaaccgtca
      601 gatcgcctgg agccaattcc aactcttttg ccttatacct actttccgta ccacttccta
      661 ccctcgaaag gcctctgagg ccaccatgaa cgacatcttc gaggctcagc gtatcgaatg
      721 gcactcatca cgtcgtaagt ggaataagac aggtcacgca gtcagagcta taggtcggct
      781 gagctcagat ctgtacgacg atgacgataa gctcgagggg agcgtggcag tcctgcctgt
      841 ggtcgactcc accaatcagt acctgctgga tcgaatcggc gagctgaaga gtggggatgc
      901 ttgcattgca gaatatcagc aggcagggag aggaagcaga gggaggaaat ggttctctcc
      961 ttttggagct aacctgtacc tgagtatgtt ttggcgcctg aagcggggac cagcagcaat
     1021 cggcctgggc ccggtcatcg gaattgtcat ggcagaagcg ctgcgaaagc tgggagcaga
     1081 caaggtgcga gtcaaatggc ccaatgacct gtatctgcag gatagaaagc tggcaggcat
     1141 cctggtggag ctggccggaa taacaggcga tgctgcacag atcgtcattg gcgccgggat
     1201 taacgtggct atgaggcgcg tggaggaaag cgtggtcaat cagggctgga tcacactgca
     1261 ggaagcaggg attaacctgg acaggaatac tctggccgct acgctgatcc gagagctgcg
     1321 ggcagccctg gaactgttcg agcaggaagg cctggctcca tatctgccac ggtgggagaa
     1381 gctggataac ttcatcaata gacccgtgaa gctgatcatt ggggacaaag agattttcgg
     1441 gattagccgg gggattgata aacagggagc cctgctgctg gaacaggacg gagttatcaa
     1501 accctggatg ggcggagaaa tcagtctgcg gtctgccgaa aagctgcagg gcggtaccgg
     1561 agggagcaaa gacaatactg tgcctctgaa gctgatcgct ctcctggcta atggcgagtt
     1621 ccatagtggc gaacagctgg gagaaaccct gggcatgtcc agggccgcta tcaacaagca
     1681 cattcagact ctgcgcgact ggggcgtgga cgtgttcacc gtgcccggaa agggctactc
     1741 tctgcccgag cctatcccgc tgctgaacgc taaacagatt ctgggacagc tggacggcct
     1801 gccggaccaa ctgactgaag agcagatcgc agaatttaaa gaggctttct ccctatttga
     1861 caaggacggg gatgggacaa taacaaccaa ggagctgggg acggtgatgc ggtctctggg
     1921 gcagaacccc acagaagcag agctgcagga catgatcaat gaagtagatg ccgacggtga
     1981 cggcacaatc gacttccctg agttcctgac aatgatggca agaaaaatga aatacaggga
     2041 cacggaagaa gaaattagag aagcgttcgg tgtgtttgat aaggatggca atggctacat
     2101 cagtgcagca gagcttcgcc acgtgatgac aaaccttgga gagaagttaa cagatgaaga
     2161 ggttgatgaa atgatcaggg aagcagacat cgatggggat ggtcaggtaa actacgaaga
     2221 gtttgtacaa atgatgacag cgaagggcaa gcccatcccc aaccccctgc tgggcctgga
     2281 cagcacctga ggcctgtcag gccaagcttc catcgataga catgataaga tacattgatg
     2341 agtttggaca aaccacaaca agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     2401 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     2461 gcattcattt tatgtttcag gttcaggggg agatgtggga ggttttttaa agcaagtaaa
     2521 acctctacaa atgtggtacc taggcgagac cctgtctcac aaaataaagt aagcccggac
     2581 tgagtgcgga aaggcgggcc tggcgggtct ggtctcccca tgcgggccac cagaggccct
     2641 gcagccttca gtcgcttgaa ggggtaatgg cgcttccact cacaaacatg gcggacagag
     2701 cgtgtgaacg agatgaacag cccctcaaaa atatggccgc cgaggctgga cggccgtgcc
     2761 ccagcagcac cgcctccgcg ccccacgtga tctctcgccg ggcacagcgc tgaccgcgga
     2821 ggtccaaccg gaagaatgtc cggattggac attcggaaga gggcccgcct tccctgggga
     2881 atctctgcgc acgcgcagaa cgcttcgacc aatgaaaaca caggaagccg tccgcgcaac
     2941 cgcgttgcgt cacttctgcc gcccctgttt caaggtatat agccgtagac ggaacttcgc
     3001 ctttctctcg gccttagcgc catttttttg ggtgagtgtt ttttggttcc tgcgttggga
     3061 ttccgtgtac aatccataga catctgacct cggcacttag catcatcaca gcaaactaac
     3121 tgtagccttt ctctctttcc ctgtagaaac ctctgcacct gaggccacca tgtctagact
     3181 ggacaagagc aaagtcataa acggcgctct ggaattactc aatggagtcg gtatcgaagg
     3241 cctgacgaca aggaaactcg ctcaaaagct gggagttgag cagcctaccc tgtactggca
     3301 cgtgaagaac aagcgggccc tgctcgatgc cctgccaatc gagatgctgg acaggcatca
     3361 tacccacttc tgccccctgg aaggcgagtc atggcaagac tttctgcgga acaacgccaa
     3421 gtcattccgc tgtgctctcc tctcacatcg cgacggggct aaagtgcatc tcggcacccg
     3481 cccaacagag aaacagtacg aaaccctgga aaatcagctc gcgttcctgt gtcagcaagg
     3541 cttctccctg gagaacgcac tgtacgctct gtccgccgtg ggccacttta cactgggctg
     3601 cgtattggag gaacaggagc atcaagtagc aaaagaggaa agagagacac ctaccaccga
     3661 ttctatgccc ccacttctga gacaagcaat tgagctgttc gaccggcagg gagccgaacc
     3721 tgccttcctt ttcggcctgg aactaatcat atgtggcctg gagaaacagc taaagtgcga
     3781 aagcggcggg ccggccgacg cccttgacga ttttgactta gacatgctcc cagccgatgc
     3841 ccttgacgac tttgaccttg atatgctgcc tgctgacgct cttgacgatt ttgaccttga
     3901 catgctcccc gggtccggcg ctactaactt cagcctgctg aagcaggctg gtgacgtcga
     3961 ggagaatcct ggtcccatga aaaagcctga actcaccgcg acgtctgtcg agaagtttct
     4021 gatcgaaaag ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg
     4081 tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga
     4141 tggtttctac aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc
     4201 ggaagtgctt gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc
     4261 acagggtgtc acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt
     4321 cgcggaggcc atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc
     4381 attcggaccg caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc
     4441 tgatccccat gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc
     4501 gcaggctctc gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt
     4561 gcacgcggat ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat
     4621 tgactggagc gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg
     4681 gaggccgtgg ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga
     4741 gcttgcagga tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta
     4801 tcagagcttg gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc
     4861 aatcgtccga tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc
     4921 cgtctggacc gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac
     4981 tcgtccgagg gcaaaggaat agttcgaagg cctgtcgtga agcttgggga tcaattctct
     5041 agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     5101 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     5161 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     5221 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     5281 tctatggctt ctgaggcgga aagaaccagc tgggcctact agctactcgg gaccccttac
     5341 cgaaacatcg ccgcattctg cagaggagtc gagtgtatgt aaacttctga cccactggga
     5401 atgtgatgaa agaaataaaa gctgaaatga atcattctct ctactattat tctgatattt
     5461 cacattctta aaataaagtg gtgatcctaa ctgacctaag acagggaatt tttactagga
     5521 ttaaatgtca ggaattgtga aaaagtgagt ttaaatgtat ttggctaagg tgtatgtaaa
     5581 cttccgactt caactgtata gggatccgct tcctcgctca ctgactcgct gcgctcggtc
     5641 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5701 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5761 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5821 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5881 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5941 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6001 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6061 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6121 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6181 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6241 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6301 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6361 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6421 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6481 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6541 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6601 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6661 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6721 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6781 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6841 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6901 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6961 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7021 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7081 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7141 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7201 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7261 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7321 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7381 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7441 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7501 gttccgcgca catttccccg aaaagtgcca cctgatgcgg tgtgaaatac cgcacagatg
     7561 cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta atattttgtt aaaatt
//
