LOCUS       sequence-11411-e        5687 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 5687)
  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 5687)
  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-2
FEATURES             Location/Qualifiers
     source          1..5687
                     /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(1215..1835,1836..2222)
                     /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(1288..1307)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     intron          2259..2831
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     complement(2963..2982)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3028..3083
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3082..3101)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3480..3497)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    complement(3922..4017)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4127..4146)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4138..4947
                     /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(4205..4224)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5037..5625
                     /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 agggtaccag tgaaagtctg caatgaattc gagttggcgc gcctgtcatt ctaaatctct
      601 ctttcagcct aaagcttttt ccccgtatcc ccccaggtgt ctgcaggctc aaagagcagc
      661 gagaagcgtt cagaggaaag cgatcccgtg ccaccttccc cgtgcccggg ctgtccccgc
      721 acgctgccgg ctcggggatg cggggggagc gccggaccgg agcggagccc cgggcggctc
      781 gctgctgccc cctagcgggg gagggacgta attacatccc tgggggcttt gggagggggc
      841 tgtccccgtg agctcccagg cgcgcctgtc attctaaatc tctctttcag cctaaagctt
      901 tttccccgta tccccccagg tgtctgcagg ctcaaagagc agcgagaagc gttcagagga
      961 aagcgatccc gtgccacctt ccccgtgccc gggctgtccc cgcacgctgc cggctcgggg
     1021 atgcgggggg agcgccggac cggagcggag ccccgggcgg ctcgctgctg ccccctagcg
     1081 ggggagggac gtaattacat ccctgggggc tttggggggg ggctgtcccc gtgagctccc
     1141 aggcgcgcca actcggtacc cggggatcca catatgttaa ttaacctgca ggtcgacgga
     1201 tccggggaat tcatatgtct agattagata aaagtaaagt gattaacagc gcattagagc
     1261 tgcttaatga ggtcggaatc gaaggtttaa caacccgtaa actcgcccag aagctaggtg
     1321 tagagcagcc tacattgtat tggcatgtaa aaaataagcg ggctttgctc gacgccttag
     1381 ccattgagat gttagatagg caccatactc acttttgccc tttagaaggg gaaagctggc
     1441 aagatttttt acgtaataac gctaaaagtt ttagatgtgc tttactaagt catcgcgatg
     1501 gagcaaaagt acatttaggt acacggccta cagaaaaaca gtatgaaact ctcgaaaatc
     1561 aattagcctt tttatgccaa caaggttttt cactagagaa tgcattatat gcactcagcg
     1621 ctgtggggca ttttacttta ggttgcgtat tggaagatca agagcatcaa gtcgctaaag
     1681 aagaaaggga aacacctact actgatagta tgccgccatt attacgacaa gctatcgaat
     1741 tatttgatca ccaaggtgca gagccagcct tcttattcgg ccttgaattg atcatatgcg
     1801 gattagaaaa acaacttaaa tgtgaaagtg ggtccgcgta cagccgcgcg cgtacgaaaa
     1861 acaattacgg gtctaccatc gagggcctgc tcgatctccc ggacgacgac gcccccgaag
     1921 aggcggggct ggcggctccg cgcctgtcct ttctccccgc gggacacacg cgcagactgt
     1981 cgacggcccc cccgaccgat gtcagcctgg gggacgagct ccacttagac ggcgaggacg
     2041 tggcgatggc gcatgccgac gcgctagacg atttcgatct ggacatgttg ggggacgggg
     2101 attccccggg tccgggattt accccccacg actccgcccc ctacggcgct ctggatatgg
     2161 ccgacttcga gtttgagcag atgtttaccg atgcccttgg aattgacgag tacggtgggt
     2221 agggggcgcg aggatcctct agagctgaga acttcagggt gagtttgggg acccttgatt
     2281 gttctttctt tttcgctatt gtaaaattca tgttatatgg agggggcaaa gttttcaggg
     2341 tgttgtttag aatgggaaga tgtcccttgt atcaccatgg accctcatga taattttgtt
     2401 tctttcactt tctactctgt tgacaaccat tgtctcctct tattttcttt tcattttctg
     2461 taactttttc gttaaacttt agcttgcatt tgtaacgaat ttttaaattc acttttgttt
     2521 atttgtcaga ttgtaagtac tttctctaat cacttttttt tcaaggcaat cagggtatat
     2581 tatattgtac ttcagcacag ttttagagaa caattgttat aattaaatga taaggtagaa
     2641 tatttctgca tataaattct ggctggcgtg gaaatattct tattggtaga aacaactaca
     2701 ccctggtcat catcctgcct ttctctttat ggttacaatg atatacactg tttgagatga
     2761 ggataaaata ctctgagtcc aaaccgggcc cctctgctaa ccatgttcat gccttcttct
     2821 ttttcctaca gctcctgggc aacgtgctgg ttgttgtgct gtctcatcat tttggcaaag
     2881 aattcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     2941 tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga
     3001 agccccttga gcatctgact tctgggtaat aaaggaaatt tattttcatt gcaatagtgt
     3061 gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac
     3121 atcagaatga gtatttggtt tagagtttgg caacatatgc catatgctgg ctgccatgaa
     3181 caaaggttgg ctataaagag gtcatcagta tatgaaacag ccccctgctg tccattcctt
     3241 attccataga aaagccttga cttgaggtta gatttttttt atattttgtt ttgtgttatt
     3301 tttttcttta acatccctaa aattttcctt acatgtttta ctagccagat ttttcctcct
     3361 ctcctgacta ctcccagtca tagctgtccc tcttctctta tggagatccc tcgactgcat
     3421 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3481 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3541 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3601 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3661 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3721 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3781 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3841 tctcaattag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3901 gctgtgcgcg gccgcagatc tcaactttat tatacaaagt tggcattata aaaaagcatt
     3961 gcttatcaat ttgttgcaac gaacaggtca ctatcagtca aaataaaatc attatttgat
     4021 atcctgcagc tctggcccgt gtctcaaaat ctctgatgtt acattgcaca agataaaaat
     4081 atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg
     4141 agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga tgctgattta
     4201 tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat ctatcgcttg
     4261 tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag cgttgccaat
     4321 gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc tcttccgacc
     4381 atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc gatccccgga
     4441 aaaacagcat tccaggtatt agaagaatat cctgattcag gtgaaaatat tgttgatgcg
     4501 ctggcagtgt tcctgcgccg gttgcattcg attcctgttt gtaattgtcc ttttaacagc
     4561 gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg
     4621 agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa agaaatgcat
     4681 aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc acttgataac
     4741 cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt cggaatcgca
     4801 gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc tccttcatta
     4861 cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa attgcagttt
     4921 catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta acattattca
     4981 gattgggccc cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     5041 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     5101 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     5161 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     5221 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     5281 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     5341 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     5401 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     5461 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     5521 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     5581 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5641 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtt
//
