LOCUS       sequence-145904-        7831 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tet-inducible human NANOG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7831)
  AUTHORS   Takashima Y, Guo G, Loos R, Nichols J, Ficz G, Krueger F, Oxley D, 
            Santos F, Clarke J, Mansfield W, Reik W, Bertone P, Smith A
  TITLE     Resetting Transcription Factor Control Circuitry toward Ground-State
            Pluripotency in Human.
  JOURNAL   Cell. 2014 Sep 11;158(6):1254-69. doi: 10.1016/j.cell.2014.08.029.
  PUBMED    25215486
REFERENCE   2  (bases 1 to 7831)
  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: 
            "10.1016/j.cell.2014.08"; journalName: "Cell"; date: "2014-09-11- 
            11"; volume: "158"; issue: "6"; pages: "1254-69"
COMMENT     Sequence Label: pPB-hCMV1-hNANOG-IV
FEATURES             Location/Qualifiers
     source          1..7831
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     623..642
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        623..641
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   660..694
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    join(1075..1093,1094..1116,1117..1135,1136..1158,1159..1177,
                     1178..1200,1201..1219,1220..1242,1243..1261,1262..1284,
                     1285..1303,1304..1326,1327..1345)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        1378..1416
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1413..1437
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          1539..2111
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     protein_bind    2176..2200
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3125..3149)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3187..3213
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    join(3249..3822,3823..3825,3826..3834)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3416..3433)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3643..3662
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3835..3837,3838..3840,3841..4554)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3880..3901)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4141..4160)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4488..4509
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4939..4956)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4945..5169
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5216..5232)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5240..5256
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5264..5270,5271..5288,5289..5294))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5309..5330
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     repeat_region   5557..5619
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(5643..5661)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5682..5698)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5706..5722
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5730..5736,5737..5754,5755..5760))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5775..5796
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5913..5930)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6084..6672)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6164..6183)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6843..7634,7635..7703))
                     /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"
     primer_bind     7466..7485
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7704..7808)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgaat tgtaatacga ctcactatag ggcgaattgg gtaccttaac
      661 cctagaaaga tagtctgcgt aaaattgacg catgcattct tgaaatattg ctctctcttt
      721 ctaaatagcg cgaatccgtc gctgtgcatt taggacatct cagtcgccgc ttggagctcc
      781 cgtgaggcgt gcttgtcaat gcggtaagtg tcactgattt tgaactataa cgaccgcgtg
      841 agtcaaaatg acgcatgatt atcttttacg tgacttttaa gatttaactc atacgataat
      901 tatattgtta tttcatgttc tacttacgtg ataacttatt atatatatat tttcttgtta
      961 tagatatcat aatttaaaca agcaaaacca aattaagggc cagctcattc ctcccactca
     1021 tgatctatag atctggatcc gtcgacaacg gcaagcttac atcgagttta ccactcccta
     1081 tcagtgatag agaaaagtga aagtcgagtt taccactccc tatcagtgat agagaaaagt
     1141 gaaagtcgag tttaccactc cctatcagtg atagagaaaa gtgaaagtcg agtttaccac
     1201 tccctatcag tgatagagaa aagtgaaagt cgagtttacc actccctatc agtgatagag
     1261 aaaagtgaaa gtcgagttta ccactcccta tcagtgatag agaaaagtga aagtcgagtt
     1321 taccactccc tatcagtgat agagaaaagt gaaagtcgag ctcggtaccc gggtcgaggt
     1381 aggcgtgtac ggtgggaggc ctatataagc agagctcgtt tagtgaaccg tcagatcgcc
     1441 tggagacgcc atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc
     1501 cgcgggactc agacacccgg gatcctgaga acttcagggt gagtttgggg acccttgatt
     1561 gttctttctt tttcgctatt gtaaaattca tgttatatgg agggggcaaa gttttcaggg
     1621 tgttgtttag aatgggaaga tgtcccttgt atcaccatgg accctcatga taattttgtt
     1681 tctttcactt tctactctgt tgacaaccat tgtctcctct tattttcttt tcattttctg
     1741 taactttttc gttaaacttt agcttgcatt tgtaacgaat ttttaaattc acttttgttt
     1801 atttgtcaga ttgtaagtac tttctctaat cacttttttt tcaaggcaat cagggtatat
     1861 tatattgtac ttcagcacag ttttagagaa caattgttat aattaaatga taaggtagaa
     1921 tatttctgca tataaattct ggctggcgtg gaaatattct tattggtaga aacaactaca
     1981 ccctggtcat catcctgcct ttctctttat ggttacaatg atatacactg tttgagatga
     2041 ggataaaata ctctgagtcc aaaccgggcc cctctgctaa ccatgttcat gccttcttct
     2101 ctttcctaca gctcctgggc aacgtgctgg ttgttgtgct gtctcatcat tttggcaaag
     2161 aattcctcga atcaaacaag tttgtacaaa aaagcaggct tcaccatgag tgtggatcca
     2221 gcttgtcccc aaagcttgcc ttgctttgaa gcatccgact gtaaagaatc ttcacctatg
     2281 cctgtgattt gtgggcctga agaaaactat ccatccttgc aaatgtcttc tgctgagatg
     2341 cctcacacgg agactgtctc tcctcttccc tcctccatgg atctgcttat tcaggacagc
     2401 cctgattctt ccaccagtcc caaaggcaaa caacccactt ctgcagagaa tagtgtcgca
     2461 aaaaaggaag acaaggtccc agtcaagaaa cagaagacca gaactgtgtt ctcttccacc
     2521 cagctgtgtg tactcaatga tagatttcag agacagaaat acctcagcct ccagcagatg
     2581 caagaactct ccaacatcct gaacctcagc tacaaacagg tgaagacctg gttccagaac
     2641 cagagaatga aatctaagag gtggcagaaa aacaactggc cgaagaatag caatggtgtg
     2701 acgcagaagg cctcagcacc tacctacccc agcctctact cttcctacca ccagggatgc
     2761 ctggtgaacc cgactgggaa ccttccaatg tggagcaacc agacctggaa caattcaacc
     2821 tggagcaacc agacccagaa catccagtcc tggagcaacc actcctggaa cactcagacc
     2881 tggtgcaccc aatcctggaa caatcaggcc tggaacagtc ccttctataa ctgtggagag
     2941 gaatctctgc agtcctgcat gcagttccag ccaaattctc ctgccagtga cttggaggct
     3001 gctttggaag ctgctgggga aggccttaat gtaatacagc agaccactag gtattttagt
     3061 actccacaaa ccatggattt attcctaaac tactccatga acatgcaacc tgaagacgtg
     3121 tgagacccag ctttcttgta caaagtggtt cgattcgaga tctcacgcgt gactagtgcc
     3181 atggcctacc catacgacgt gccagactac gcctagcggc cgctcgataa gcttgatatc
     3241 gaattccgcc cctctccctc ccccccccct aacgttactg gccgaagccg cttggaataa
     3301 ggccggtgtg cgtttgtcta tatgttattt tccaccatat tgccgtcttt tggcaatgtg
     3361 agggcccgga aacctggccc tgtcttcttg acgagcattc ctaggggtct ttcccctctc
     3421 gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct ggaagcttct
     3481 tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc ggaacccccc acctggcgac
     3541 aggtgcctct gcggccaaaa gccacgtgta taagatacac ctgcaaaggc ggcacaaccc
     3601 cagtgccacg ttgtgagttg gatagttgtg gaaagagtca aatggctctc ctcaagcgta
     3661 ttcaacaagg ggctgaagga tgcccagaag gtaccccatt gtatgggatc tgatctgggg
     3721 cctcggtgca catgctttac atgtgtttag tcgaggttaa aaaacgtcta ggccccccga
     3781 accacgggga cgtggttttc ctttgaaaaa cacgatgata atatggccac aaccatggtg
     3841 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     3901 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     3961 ctgaccctga agctgatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     4021 accaccctgg gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac
     4081 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     4141 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     4201 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     4261 gagtacaact acaacagcca caacgtctat atcaccgccg acaagcagaa gaacggcatc
     4321 aaggccaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac
     4381 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     4441 agctaccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     4501 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaagcggc
     4561 cagcttatcg actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg
     4621 aatagtcgag aaattgatga tctattaaac aataaagatg tccacatgga agtttttcct
     4681 gtcatacttt gttaagaagg gtgagaacag agtacctaca ttttgaatgg aaggattgga
     4741 gctacggggg tgggggtggg gtgggattag ataaatgcct gctctttact gaaggctctt
     4801 tactattgct ttatgataat gtttcatagt tggatatcat aatttaaaca agcaaaacca
     4861 aattaagggc cagctcattc ctcccactca tgatctatag atcctaagct tatcgatgat
     4921 ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     4981 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     5041 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     5101 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     5161 gctctatggc ttctgaggcg gaaagaacct gcagcccaag cttggcgtaa tcatggtcat
     5221 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5281 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5341 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gcggatcctt tgttacttta
     5401 tagaagaaat tttgagtttt tgtttttttt taataaataa ataaacataa ataaattgtt
     5461 tgttgaattt attattagta tgtaagtgta aatataataa aacttaatat ctattcaaat
     5521 taataaataa acctcgatat acagaccgat aaaacacatg cgtcaatttt acgcatgatt
     5581 atctttaacg tacgtcacaa tatgattatc tttctagggt taaagagctc cagcttttgt
     5641 tccctttagt gagggttaat ttcgagcttg gcgtaatcat ggtcatagct gtttcctgtg
     5701 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5761 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5821 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5881 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5941 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     6001 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     6061 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     6121 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     6181 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6241 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6301 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6361 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6421 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6481 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     6541 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6601 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6661 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6721 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6781 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6841 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6901 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6961 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     7021 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     7081 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     7141 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7201 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     7261 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7321 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7381 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7441 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7501 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7561 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7621 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7681 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7741 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7801 gttccgcgca catttccccg aaaagtgcca c
//
