LOCUS Exported 8988 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Over-expression of human KLF4. ACCESSION . VERSION . KEYWORDS pPB-PGK-hKLF4 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8988) 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 8988) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8988 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind complement(1430..1454) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" primer_bind complement(1475..1491) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind 1499..1515 /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(1523..1553) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1568..1589 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(1649..1666) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 1655..1879 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" protein_bind complement(1956..1989) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(2212..2231) /label=T7 /note="T7 promoter, forward primer" promoter complement(2213..2231) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2241..2258) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2241..2257) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2250..2272) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 2399..2854 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2486..2505) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2696..2717 /label=F1ori-F /note="F1 origin, forward primer" promoter 2880..2984 /gene="bla" /label=AmpR promoter CDS 2985..3845 /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 complement(3203..3222) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4016..4604 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4505..4524 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 4758..4775 /label=L4440 /note="L4440 vector, forward primer" protein_bind 4892..4913 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 4928..4958 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4966..4982 /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)." primer_bind 4990..5006 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5025..5045 /label=T3 /note="T3 promoter, forward primer" promoter 5027..5045 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" repeat_region 5068..5102 /label=piggyBac left (5') inverted repeat /note="piggyBac transposon-specific inverted terminal repeat sequence (ITR)" CDS complement(5585..6610) /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" promoter complement(6629..7128) /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" primer_bind 7070..7093 /label=MSCV-rev /note="Murine stem cell virus, reverse primer" protein_bind complement(7151..7184) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." enhancer 7200..7579 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 7582..7857 /label=chicken beta-actin promoter intron 7858..8866 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 8790..8812 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" protein_bind 8944..8968 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" ORIGIN 1 atggctgtca gtgacgcgct gctcccatct ttctccacgt tcgcgtctgg cccggcggga 61 agggagaaga cactgcgtca agcaggtgcc ccgaataacc gctggcggga ggagctctcc 121 cacatgaagc gacttccccc agtgcttccc ggccgcccct atgacctggc ggcggcgacc 181 gtggccacag acctggagag cggcggagcc ggtgcggctt gcggcggtag caacctggcg 241 cccctacctc ggagagagac cgaggagttc aacgatctcc tggacctgga ctttattctc 301 tccaattcgc tgacccatcc tccggagtca gtggccgcca ccgtgtcctc gtcagcgtca 361 gcctcctctt cgtcgtcgcc gtcgagcagc ggccctgcca gcgcgccctc cacctgcagc 421 ttcacctatc cgatccgggc cgggaacgac ccgggcgtgg cgccgggcgg cacgggcgga 481 ggcctcctct atggcaggga gtccgctccc cctccgacgg ctcccttcaa cctggcggac 541 atcaacgacg tgagcccctc gggcggcttc gtggccgagc tcctgcggcc agaattggac 601 ccggtgtaca ttccgccgca gcagccgcag ccgccaggtg gcgggctgat gggcaagttc 661 gtgctgaagg cgtcgctgag cgcccctggc agcgagtacg gcagcccgtc ggtcatcagc 721 gtcagcaaag gcagccctga cggcagccac ccggtggtgg tggcgcccta caacggcggg 781 ccgccgcgca cgtgccccaa gatcaagcag gaggcggtct cttcgtgcac ccacttgggc 841 gctggacccc ctctcagcaa tggccaccgg ccggctgcac acgacttccc cctggggcgg 901 cagctcccca gcaggactac cccgaccctg ggtcttgagg aagtgctgag cagcagggac 961 tgtcaccctg ccctgccgct tcctcccggc ttccatcccc acccggggcc caattaccca 1021 tccttcctgc ccgatcagat gcagccgcaa gtcccgccgc tccattacca agagctcatg 1081 ccacccggtt cctgcatgcc agaggagccc aagccaaaga ggggaagacg atcgtggccc 1141 cggaaaagga ccgccaccca cacttgtgat tacgcgggct gcggcaaaac ctacacaaag 1201 agttcccatc tcaaggcaca cctgcgaacc cacacaggtg agaaacctta ccactgtgac 1261 tgggacggct gtggatggaa attcgcccgc tcagatgaac tgaccaggca ctaccgtaaa 1321 cacacggggc accgcccgtt ccagtgccaa aaatgcgacc gagcattttc caggtcggac 1381 cacctcgcct tacacatgaa gagacatttt taaaagggtg ggcgcgccga cccagctttc 1441 ttgtacaaag tggtgatagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt 1501 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg 1561 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt 1621 cgggaaacct gtcgtgccag cggatcagcc tcgactgtgc cttctagttg ccagccatct 1681 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt 1741 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg 1801 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg 1861 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc gagatccact 1921 agttctagcc tcgaggctag agcggccgcg tcgagataac ttcgtataat gtatgctata 1981 cgaagttatg ttttgttact ttatagaaga aattttgagt ttttgttttt ttttaataaa 2041 taaataaaca taaataaatt gtttgttgaa tttattatta gtatgtaagt gtaaatataa 2101 taaaacttaa tatctattca aattaataaa taaacgtcaa ttttacgcat gattatcttt 2161 aacgtacgtc acaatatgat tatctttcta gggttaaaga gctccaattc gccctatagt 2221 gagtcgtatt acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct 2281 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc 2341 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac 2401 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct 2461 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg 2521 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt 2581 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca 2641 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga 2701 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa 2761 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac 2821 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc 2881 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac 2941 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt 3001 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag 3061 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg 3121 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa 3181 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc 3241 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag 3301 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa 3361 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc 3421 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg 3481 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa 3541 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa 3601 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg 3661 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag 3721 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg 3781 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt 3841 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt 3901 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac 3961 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag 4021 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg 4081 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca 4141 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga 4201 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca 4261 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc 4321 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca 4381 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa 4441 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc 4501 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc 4561 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg 4621 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat 4681 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca 4741 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca 4801 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg 4861 actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac 4921 cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac 4981 aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt aaccctcact 5041 aaagggaaca aaagctgggt accttaaccc tagaaagata gtctgcgtaa aattgacgca 5101 tgcattcttg aaatattgct ctctctttct aaatagcgcg aatccgtcgc tgtgcattta 5161 ggacatctca gtcgccgctt ggagctcccg tgaggcgtgc ttgtcaatgc ggtaagtgtc 5221 actgattttg aactataacg accgcgtgag tcaaaatgac gcatgattat cttttacgtg 5281 acttttaaga tttaactcat acgataatta tattgttatt tcatgttcta cttacgtgat 5341 aacttattat atatatattt tcttgttata gatatccaac tatgaaacat tatcataaag 5401 caatagtaaa gagccttcag taaagagcag gcatttatct aatcccaccc cacccccacc 5461 cccgtagctc caatccttcc attcaaaatg taggtactct gttctcaccc ttcttaacaa 5521 agtatgacag gaaaaaactt ccatgtggac atctttattg tttaatagat catcaatttc 5581 tcgactattc ctttgccctc ggacgagtgc tggggcgtcg gtttccacta tcggcgagta 5641 cttctacaca gccatcggtc cagacggccg cgcttctgcg ggcgatttgt gtacgcccga 5701 cagtcccggc tccggatcgg acgattgcgt cgcatcgacc ctgcgcccaa gctgcatcat 5761 cgaaattgcc gtcaaccaag ctctgataga gttggtcaag accaatgcgg agcatatacg 5821 cccggagccg cggcgatcct gcaagctccg gatgcctccg ctcgaagtag cgcgtctgct 5881 gctccataca agccaaccac ggcctccaga agaagatgtt ggcgacctcg tattgggaat 5941 ccccgaacat cgcctcgctc cagtcaatga ccgctgttat gcggccattg tccgtcagga 6001 cattgttgga gccgaaatcc gcgtgcacga ggtgccggac ttcggggcag tcctcggccc 6061 aaagcatcag ctcatcgaga gcctgcgcga cggacgcact gacggtgtcg tccatcacag 6121 tttgccagtg atacacatgg ggatcagcaa tcgcgcatat gaaatcacgc catgtagtgt 6181 attgaccgat tccttgcggt ccgaatgggc cgaacccgct cgtctggcta agatcggccg 6241 cagcgatcgc atccatggcc tccgcgaccg gctgcagaac agcgggcagt tcggtttcag 6301 gcaggtcttg caacgtgaca ccctgtgcac ggcgggagat gcaataggtc aggctctcgc 6361 tgaattcccc aatgtcaagc acttccggaa tcgggagcgc ggccgatgca aagtgccgat 6421 aaacataacg atctttgtag aaaccatcgg cgcagctatt tacccgcagg acatatccac 6481 gccctcctac atcgaagctg aaagcacgag attcttcgcc ctccgagagc tgcatcaggt 6541 cggagacgct gtcgaacttt tcgatcagaa acttctcgac agacgtcgcg gtgagttcag 6601 gctttttcat ggtggcggct ggatcggtcg aaaggcccgg agatgaggaa gaggagaaca 6661 gcgcggcaga cgtgcgcttt tgaagcgtgc gagaatgccg ggcctccgga ggaccttcgg 6721 gcgcccgccc cgcccctgag cccgcccctg agcccgcccc cggacccacc ccttcccagc 6781 ctctgagccc agaaagcgaa ggagcaaagc tgctattggc cgctgcccca aaggcctacc 6841 cgcttccatt gctcagcggt gctgtccatc tgcacgagac tagtgagacg tgctacttcc 6901 atttgtcacg tcctgcacga cgcgagctgc ggggcggggg ggaacttcct gactagggga 6961 ggagtagaag gtggcgcgaa ggggccacca aagaacggag ccggttggcg cctaccggtg 7021 gatgtggaat gtgtgcgagg ccagaggcca cttgtgtagc gccaagtgcc agcggggctg 7081 ctaaagcgca tgctccagac tgccttggga aaagcgcctc ccctacccgg tagaattcat 7141 cgatgatatc ataacttcgt ataatgtatg ctatacgaag ttatgcggcc ctcgaggtcg 7201 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc 7261 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa 7321 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac 7381 tttccattga cgtcaatggg tggactattt acggtaaact gcccacttgg cagtacatca 7441 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg 7501 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt 7561 agtcatcgct attaccatgg gtcgaggtga gccccacgtt ctgcttcact ctccccatct 7621 cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga 7681 tgggggcggg gggggggggg gcgcgcgcca ggcggggcgg ggcggggcga ggggcggggc 7741 ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct 7801 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga 7861 gtcgctgcgt tgccttcgcc ccgtgccccg ctccgcgccg cctcgcgccg cccgccccgg 7921 ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc 7981 tgtaattagc gcttggttta atgacggctc gtttcttttc tgtggctgcg tgaaagcctt 8041 aaagggctcc gggagggccc tttgtgcggg ggggagcggc tcggggggtg cgtgcgtgtg 8101 tgtgtgcgtg gggagcgccg cgtgcggccc gcgctgcccg gcggctgtga gcgctgcggg 8161 cgcggcgcgg ggctttgtgc gctccgcgtg tgcgcgaggg gagcgcggcc gggggcggtg 8221 ccccgcggtg cgggggggct gcgaggggaa caaaggctgc gtgcggggtg tgtgcgtggg 8281 ggggtgagca gggggtgtgg gcgcggcggt cgggctgtaa cccccccctg cacccccctc 8341 cccgagttgc tgagcacggc ccggcttcgg gtgcggggct ccgtgcgggg cgtggcgcgg 8401 ggctcgccgt gccgggcggg gggtggcggc aggtgggggt gccgggcggg gcggggccgc 8461 ctcgggccgg ggagggctcg ggggaggggc gcggcggccc cggagcgccg gcggctgtcg 8521 aggcgcggcg agccgcagcc attgcctttt atggtaatcg tgcgagaggg cgcagggact 8581 tcctttgtcc caaatctggc ggagccgaaa tctgggaggc gccgccgcac cccctctagc 8641 gggcgcgggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag ggccttcgtg 8701 cgtcgccgcg ccgccgtccc cttctccatc tccagcctcg gggctgccgc agggggacgg 8761 ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga ccggcggctc 8821 tagagcctct gctaaccatg ttcatgcctt cttctttttc ctacagctcc tgggcaacgt 8881 gctggttgtt gtgctgtctc atcattttgg caaagaattc ctcgacggta tcgataagct 8941 atcacaagtt tgtacaaaaa agcaggctcc gcggccgccc ccttcacc //