LOCUS       sequence-12684-e        5075 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5075)
  AUTHORS   Hsiao EC, Nguyen TD, Ng JK, Scott MJ, Chang WC, Zahed H, Conklin BR
  TITLE     Constitutive Gs activation using a single-construct 
            tetracycline-inducible expression system in embryonic stem cells and
            mice.
  JOURNAL   Stem Cell Res Ther. 2011 Mar 4. 2(2):11.
  PUBMED    21375737
REFERENCE   2  (bases 1 to 5075)
  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: "Stem Cell 
            Res Ther. 2011 Mar 4. 2(2):11."
COMMENT     Sequence Label: pEnt L1L3 tTA-3
FEATURES             Location/Qualifiers
     source          1..5075
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      103..189
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      281..308
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     316..339
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    358..457
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(490..523)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             join(603..1223,1224..1610)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTI
                     EGLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHA
                     DALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(676..695)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     intron          1647..2219
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     complement(2351..2370)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2416..2471
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2470..2489)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2868..2885)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    complement(3310..3405)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3515..3534)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3526..4335
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3593..3612)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4425..5013
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgctagca tggatctcgg
       61 ggacgtctaa ctactaagcg agagtaggga actgccaggc atcaaataaa acgaaaggct
      121 cagtcggaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt
      181 aggacaaatc cgccgggagc ggatttgaac gttgtgaagc aacggcccgg agggtggcgg
      241 gcaggacgcc cgccataaac tgccaggcat caaactaagc agaaggccat cctgacggat
      301 ggcctttttg cgtttctaca aactcttcct gttagttagt tacttaagct cgggccccaa
      361 ataatgattt tattttgact gatagtgacc tgttcgttgc aacaaattga taagcaatgc
      421 ttttttataa tgccaacttt gtacaaaaaa gcaggctggc gccggaacca attcagtcga
      481 cgtttaaaca taacttcgta tagcatacat tatacgaagt tatctcgagc ctcaggaaac
      541 agggtacccg gggatccaca tatgttaatt aacctgcagg tcgacggatc cggggaattc
      601 atatgtctag attagataaa agtaaagtga ttaacagcgc attagagctg cttaatgagg
      661 tcggaatcga aggtttaaca acccgtaaac tcgcccagaa gctaggtgta gagcagccta
      721 cattgtattg gcatgtaaaa aataagcggg ctttgctcga cgccttagcc attgagatgt
      781 tagataggca ccatactcac ttttgccctt tagaagggga aagctggcaa gattttttac
      841 gtaataacgc taaaagtttt agatgtgctt tactaagtca tcgcgatgga gcaaaagtac
      901 atttaggtac acggcctaca gaaaaacagt atgaaactct cgaaaatcaa ttagcctttt
      961 tatgccaaca aggtttttca ctagagaatg cattatatgc actcagcgct gtggggcatt
     1021 ttactttagg ttgcgtattg gaagatcaag agcatcaagt cgctaaagaa gaaagggaaa
     1081 cacctactac tgatagtatg ccgccattat tacgacaagc tatcgaatta tttgatcacc
     1141 aaggtgcaga gccagccttc ttattcggcc ttgaattgat catatgcgga ttagaaaaac
     1201 aacttaaatg tgaaagtggg tccgcgtaca gccgcgcgcg tacgaaaaac aattacgggt
     1261 ctaccatcga gggcctgctc gatctcccgg acgacgacgc ccccgaagag gcggggctgg
     1321 cggctccgcg cctgtccttt ctccccgcgg gacacacgcg cagactgtcg acggcccccc
     1381 cgaccgatgt cagcctgggg gacgagctcc acttagacgg cgaggacgtg gcgatggcgc
     1441 atgccgacgc gctagacgat ttcgatctgg acatgttggg ggacggggat tccccgggtc
     1501 cgggatttac cccccacgac tccgccccct acggcgctct ggatatggcc gacttcgagt
     1561 ttgagcagat gtttaccgat gcccttggaa ttgacgagta cggtgggtag ggggcgcgag
     1621 gatcctctag agctgagaac ttcagggtga gtttggggac ccttgattgt tctttctttt
     1681 tcgctattgt aaaattcatg ttatatggag ggggcaaagt tttcagggtg ttgtttagaa
     1741 tgggaagatg tcccttgtat caccatggac cctcatgata attttgtttc tttcactttc
     1801 tactctgttg acaaccattg tctcctctta ttttcttttc attttctgta actttttcgt
     1861 taaactttag cttgcatttg taacgaattt ttaaattcac ttttgtttat ttgtcagatt
     1921 gtaagtactt tctctaatca cttttttttc aaggcaatca gggtatatta tattgtactt
     1981 cagcacagtt ttagagaaca attgttataa ttaaatgata aggtagaata tttctgcata
     2041 taaattctgg ctggcgtgga aatattctta ttggtagaaa caactacacc ctggtcatca
     2101 tcctgccttt ctctttatgg ttacaatgat atacactgtt tgagatgagg ataaaatact
     2161 ctgagtccaa accgggcccc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     2221 tcctgggcaa cgtgctggtt gttgtgctgt ctcatcattt tggcaaagaa ttcactcctc
     2281 aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc caatgccctg gctcacaaat
     2341 accactgaga tctttttccc tctgccaaaa attatgggga catcatgaag ccccttgagc
     2401 atctgacttc tgggtaataa aggaaattta ttttcattgc aatagtgtgt tggaattttt
     2461 tgtgtctctc actcggaagg acatatggga gggcaaatca tttaaaacat cagaatgagt
     2521 atttggttta gagtttggca acatatgcca tatgctggct gccatgaaca aaggttggct
     2581 ataaagaggt catcagtata tgaaacagcc ccctgctgtc cattccttat tccatagaaa
     2641 agccttgact tgaggttaga ttttttttat attttgtttt gtgttatttt tttctttaac
     2701 atccctaaaa ttttccttac atgttttact agccagattt ttcctcctct cctgactact
     2761 cccagtcata gctgtccctc ttctcttatg gagatccctc gactgcatta atgaatcggc
     2821 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     2881 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     2941 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     3001 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     3061 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     3121 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     3181 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcaattagct
     3241 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgcgcggc
     3301 cgcagatctc aactttatta tacaaagttg gcattataaa aaagcattgc ttatcaattt
     3361 gttgcaacga acaggtcact atcagtcaaa ataaaatcat tatttgatat cctgcagctc
     3421 tggcccgtgt ctcaaaatct ctgatgttac attgcacaag ataaaaatat atcatcatga
     3481 acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa
     3541 cgggaaacgt cgaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa
     3601 tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgcttgta tgggaagccc
     3661 gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat
     3721 gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt
     3781 atccgtactc ctgatgatgc atggttactc accactgcga tccccggaaa aacagcattc
     3841 caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc
     3901 ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt
     3961 cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat
     4021 gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa acttttgcca
     4081 ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac
     4141 gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag
     4201 gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt
     4261 tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc
     4321 gatgagtttt tctaatcaga attggttaat tggttgtaac attattcaga ttgggccccg
     4381 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     4441 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     4501 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     4561 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     4621 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     4681 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     4741 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     4801 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     4861 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     4921 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     4981 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     5041 ttcctggcct tttgctggcc ttttgctcac atgtt
//
