LOCUS       sequence-11412-e        5622 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 5622)
  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 5622)
  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 EF1a-tTA-3
FEATURES             Location/Qualifiers
     source          1..5622
                     /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)."
     promoter        602..813
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             826..1094
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             join(1150..1770,1771..2157)
                     /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(1223..1242)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     intron          2194..2766
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     complement(2898..2917)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2963..3018
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3017..3036)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3415..3432)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    complement(3857..3952)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4062..4081)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4073..4882
                     /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(4140..4159)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4972..5560
                     /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 aagatctgcg atcgctccgg tgcccgtcag
      601 tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt cggcaattga
      661 accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt gtactggctc
      721 cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt
      781 ctttttcgca acgggtttgc cgccagaaca cagctgaagc ttcgaggggc tcgcatctct
      841 ccttcacgcg cccgccgccc tacctgaggc cgccatccac gccggttgag tcgcgttctg
      901 ccgcctcccg cctgtggtgc ctcctgaact gcgtccgccg tctaggtaag tttaaagctc
      961 aggtcgagac cgggcctttg tccggcgctc ccttggagcc tacctagact cagccggctc
     1021 tccacgcttt gcctgaccct gcttgctcaa ctctacgtct ttgtttcgtt ttctgttctg
     1081 cgccgttaca gatccaagct gtgaccggcg cctacaaggc ctgcaggtcg acggatccgg
     1141 ggaattcata tgtctagatt agataaaagt aaagtgatta acagcgcatt agagctgctt
     1201 aatgaggtcg gaatcgaagg tttaacaacc cgtaaactcg cccagaagct aggtgtagag
     1261 cagcctacat tgtattggca tgtaaaaaat aagcgggctt tgctcgacgc cttagccatt
     1321 gagatgttag ataggcacca tactcacttt tgccctttag aaggggaaag ctggcaagat
     1381 tttttacgta ataacgctaa aagttttaga tgtgctttac taagtcatcg cgatggagca
     1441 aaagtacatt taggtacacg gcctacagaa aaacagtatg aaactctcga aaatcaatta
     1501 gcctttttat gccaacaagg tttttcacta gagaatgcat tatatgcact cagcgctgtg
     1561 gggcatttta ctttaggttg cgtattggaa gatcaagagc atcaagtcgc taaagaagaa
     1621 agggaaacac ctactactga tagtatgccg ccattattac gacaagctat cgaattattt
     1681 gatcaccaag gtgcagagcc agccttctta ttcggccttg aattgatcat atgcggatta
     1741 gaaaaacaac ttaaatgtga aagtgggtcc gcgtacagcc gcgcgcgtac gaaaaacaat
     1801 tacgggtcta ccatcgaggg cctgctcgat ctcccggacg acgacgcccc cgaagaggcg
     1861 gggctggcgg ctccgcgcct gtcctttctc cccgcgggac acacgcgcag actgtcgacg
     1921 gcccccccga ccgatgtcag cctgggggac gagctccact tagacggcga ggacgtggcg
     1981 atggcgcatg ccgacgcgct agacgatttc gatctggaca tgttggggga cggggattcc
     2041 ccgggtccgg gatttacccc ccacgactcc gccccctacg gcgctctgga tatggccgac
     2101 ttcgagtttg agcagatgtt taccgatgcc cttggaattg acgagtacgg tgggtagggg
     2161 gcgcgaggat cctctagagc tgagaacttc agggtgagtt tggggaccct tgattgttct
     2221 ttctttttcg ctattgtaaa attcatgtta tatggagggg gcaaagtttt cagggtgttg
     2281 tttagaatgg gaagatgtcc cttgtatcac catggaccct catgataatt ttgtttcttt
     2341 cactttctac tctgttgaca accattgtct cctcttattt tcttttcatt ttctgtaact
     2401 ttttcgttaa actttagctt gcatttgtaa cgaattttta aattcacttt tgtttatttg
     2461 tcagattgta agtactttct ctaatcactt ttttttcaag gcaatcaggg tatattatat
     2521 tgtacttcag cacagtttta gagaacaatt gttataatta aatgataagg tagaatattt
     2581 ctgcatataa attctggctg gcgtggaaat attcttattg gtagaaacaa ctacaccctg
     2641 gtcatcatcc tgcctttctc tttatggtta caatgatata cactgtttga gatgaggata
     2701 aaatactctg agtccaaacc gggcccctct gctaaccatg ttcatgcctt cttctttttc
     2761 ctacagctcc tgggcaacgt gctggttgtt gtgctgtctc atcattttgg caaagaattc
     2821 actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa tgccctggct
     2881 cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat catgaagccc
     2941 cttgagcatc tgacttctgg gtaataaagg aaatttattt tcattgcaat agtgtgttgg
     3001 aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt aaaacatcag
     3061 aatgagtatt tggtttagag tttggcaaca tatgccatat gctggctgcc atgaacaaag
     3121 gttggctata aagaggtcat cagtatatga aacagccccc tgctgtccat tccttattcc
     3181 atagaaaagc cttgacttga ggttagattt tttttatatt ttgttttgtg ttattttttt
     3241 ctttaacatc cctaaaattt tccttacatg ttttactagc cagatttttc ctcctctcct
     3301 gactactccc agtcatagct gtccctcttc tcttatggag atccctcgac tgcattaatg
     3361 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     3421 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     3481 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     3541 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     3601 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     3661 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3721 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3781 attagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3841 gcgcggccgc agatctcaac tttattatac aaagttggca ttataaaaaa gcattgctta
     3901 tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata aaatcattat ttgatatcct
     3961 gcagctctgg cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc
     4021 atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     4081 tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg
     4141 gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg
     4201 gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt
     4261 tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa
     4321 gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac
     4381 agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc
     4441 agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg
     4501 cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga
     4561 ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact
     4621 tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat
     4681 ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg
     4741 ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa
     4801 acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt
     4861 gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacatt attcagattg
     4921 ggccccgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4981 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5041 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     5101 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     5161 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     5221 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     5281 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     5341 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5401 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5461 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5521 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5581 tttacggttc ctggcctttt gctggccttt tgctcacatg tt
//
