LOCUS Exported 7728 bp ds-DNA circular SYN 17-JAN-2023 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pCAGGS-flpE-puro SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7728) AUTHORS Beard C, Hochedlinger K, Plath K, Wutz A, Jaenisch R TITLE Efficient method to generate single-copy transgenic mice by site-specific integration in embryonic stem cells. JOURNAL Genesis. 2006 Jan . 44(1):23-8. PUBMED 16400644 REFERENCE 2 (bases 1 to 7728) AUTHORS . TITLE Direct Submission JOURNAL Exported Jan 17, 2023 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7728 /organism="synthetic DNA construct" /mol_type="other DNA" polyA_signal 514..648 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" enhancer 662..1041 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" intron 1319..2335 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 2259..2281 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 2343..2362 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" regulatory 2396..2405 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2402..3700 /codon_start=1 /product="nuclear-targeted site-specific recombinase" /label=FLPe /note="FLPe is a fusion between the SV40 nuclear localization signal and a thermostable version of the Saccharomyces cerevisiae site-specific recombinase FLP. Recombination occurs at FRT sequences." /translation="MAPKKKRKVMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAA ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK LIPAWEFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESI WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY QLLKDNLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI" misc_feature 3739..4325 /label=IRES2 /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(3906..3923) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 4133..4152 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 4326..4925 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(4326..4345) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4822..4842 /label=Puro-F /note="Puromycin resistance gene, forward primer" primer_bind complement(5006..5023) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 5012..5236 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(5283..5299) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5283..5299) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5296..5318) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5307..5323 /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(5331..5361) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5376..5397 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5456..5651 /label=SV40 promoter /note="SV40 early promoter" rep_origin 5502..5637 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5564..5583 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 5657..5791 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5859..5876) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(6030..6618) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(6110..6129) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6789..7649) /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 7412..7431 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" ORIGIN 1 tattcagcta tattcacata gtggtgaggc tgaaatgagg cgggaagagg cggttggggc 61 ttaattatat caatttgggt ggccccacag cgcctccaag gcgccagtcc tgttttgaca 121 agttgcctct ggaagcctct cttctttttc tccagagtaa gcggaggcca ggggcccccg 181 gcctctgctt aatactaaaa aaaacagctg ttgtcatagt aatgattggg tggaaacatt 241 ccaggcctgg gtggagaggc tttttgcttc ctcttgcaaa accacactga cattccaggc 301 ctgggtggag aggctttttg cttcctcttg caaaaccaca ctgccctctg gagggcagtt 361 gcctagcaac taattaaaag aggatgtcgc acggccagct gcggtcagtt agtcacttcc 421 tgcttaactg acttgacatt ttctatttta agagtcggga ggaaaattac tgtgttggag 481 gccctccgcc atcttctgaa gctgaatcga attaacttgt ttattgcagc ttataatggt 541 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct 601 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggatctg atatcatcgt 661 cgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc 721 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc 781 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg 841 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat 901 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc 961 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta 1021 ttagtcatcg ctattaccat ggtcgaggtg agccccacgt tctgcttcac tctccccatc 1081 tcccccccct ccccaccccc aattttgtat ttatttattt tttaattatt ttgtgcagcg 1141 atgggggcgg gggggggggg ggcgcgcgcc aggggggggg gggggggggg gggggggggg 1201 gggggggggg gggggggggg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc 1261 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg 1321 agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc 1381 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc 1441 gggctgtaat tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag 1501 ccttgagggg ctccgggagg gccctttgtg cggggggagc ggctcggggg gtgcgtgcgt 1561 gtgtgtgtgc gtggggagcg ccgcgtgcgg ctccgcgctg cccggcggct gtgagcgctg 1621 cgggcgcggc gcggggcttt gtgcgctccg cagtgtgcgc gaggggagcg cggccggggg 1681 cggtgccccg cggtgcgggg ggggctgcga ggggaacaaa ggctgcgtgc ggggtgtgtg 1741 cgtggggggg tgagcagggg gtgtgggcgc gtcggtcggg ctgcaacccc ccctgcaccc 1801 ccctccccga gttgctgagc acggcccggc ttcgggtgcg gggctccgta cggggcgtgg 1861 cgcggggctc gccgtgccgg gcggggggtg gcggcaggtg ggggtgccgg gcggggcggg 1921 gccgcctcgg gccggggggg gctcgggggg ggggcgcggc ggcccccgga gcgccggcgg 1981 ctgtcgaggc gcggcgagcc gcagccattg ccttttatgg taatcgtgcg agagggcgca 2041 gggacttcct ttgtcccaaa tctgtgcgga gccgaaatct gggaggcgcc gccgcacccc 2101 ctctagcggg cgcggggcga agcggtgcgg cgccggcagg aaggaaatgg gcggggaggg 2161 ccttcgtgcg tcgccgcgcc gccgtcccct tctccctctc cagcctcggg gctgtccgcg 2221 gggggacggc tgccttcggg ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac 2281 cggcggctct agagcctctg ctaaccatgt tcatgccttc ttctttttcc tacagctcct 2341 gggcaacgtg ctggttattg tgctgtctca tcattttggc aaagaattac ccgccgccac 2401 catggctccc aagaagaaga ggaaggtgat gagtcaattt gatatattat gtaaaacacc 2461 acctaaggtc ctggttcgtc agtttgtgga aaggtttgaa agaccttcag gggaaaaaat 2521 agcatcatgt gctgctgaac taacctattt atgttggatg attactcata acggaacagc 2581 aatcaagaga gccacattca tgagctataa tactatcata agcaattcgc tgagtttcga 2641 tattgtcaac aaatcactcc agtttaaata caagacgcaa aaagcaacaa ttctggaagc 2701 ctcattaaag aaattaattc ctgcttggga atttacaatt attccttaca atggacaaaa 2761 acatcaatct gatatcactg atattgtaag tagtttgcaa ttacagttcg aatcatcgga 2821 agaagcagat aagggaaata gccacagtaa aaaaatgctt aaagcacttc taagtgaggg 2881 tgaaagcatc tgggagatca ctgagaaaat actaaattcg tttgagtata cctcgagatt 2941 tacaaaaaca aaaactttat accaattcct cttcctagct actttcatca attgtggaag 3001 attcagcgat attaagaacg ttgatccgaa atcatttaaa ttagtccaaa ataagtatct 3061 gggagtaata atccagtgtt tagtgacaga gacaaagaca agcgttagta ggcacatata 3121 cttctttagc gcaaggggta ggatcgatcc acttgtatat ttggatgaat ttttgaggaa 3181 ttctgaacca gtcctaaaac gagtaaatag gaccggcaat tcttcaagca acaaacagga 3241 ataccaatta ttaaaagata acttagtcag atcgtacaac aaggctttga agaaaaatgc 3301 gccttatcca atctttgcta taaagaatgg cccaaaatct cacattggaa gacatttgat 3361 gacctcattt ctgtcaatga agggcctaac ggagttgact aatgttgtgg gaaattggag 3421 cgataagcgt gcttctgccg tggccaggac aacgtatact catcagataa cagcaatacc 3481 tgatcactac ttcgcactag tttctcggta ctatgcatat gatccaatat caaaggaaat 3541 gatagcattg aaggatgaga ctaatccaat tgaggagtgg cagcatatag aacagctaaa 3601 gggtagtgct gaaggaagca tacgataccc cgcatggaat gggataatat cacaggaggt 3661 actagactac ctttcatcct acataaatag acgcatataa tgatctagag tcgagttaat 3721 taagaattcc gccccccccc cctctccctc ccccccccct aacgttactg gccgaagccg 3781 cttggaataa ggccggtgtg cgtttgtcta tatgttattt tccaccatat tgccgtcttt 3841 tggcaatgtg agggcccgga aacctggccc tgtcttcttg acgagcattc ctaggggtct 3901 ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct 3961 ggaagcttct tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc ggaacccccc 4021 acctggcgac aggtgcctct gcggccaaaa gccacgtgta taagatacac ctgcaaaggc 4081 ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg gaaagagtca aatggctctc 4141 ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag gtaccccatt gtatgggatc 4201 tgatctgggg cctcggtgca catgctttac atgtgtttag tcgaggttaa aaaaacgtct 4261 aggccccccg aaccacgggg acgtggtttt cctttgaaaa acacgatgat aatatggcca 4321 caaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtccccaggg 4381 ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgatc 4441 cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc 4501 tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc 4561 cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga 4621 gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca 4681 aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc 4741 tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct 4801 tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca 4861 ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg 4921 cctgacgccc gccccacgac ccgcagcgcc cgaccgaaag gagcgcacga ccccatgcat 4981 cgatgatcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt 5041 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc 5101 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt 5161 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat 5221 gcggtgggct ctatggcttc tgaggcggaa agaacctgca gcccaagctt ggcgtaatca 5281 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga 5341 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt 5401 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg gatccgcatc 5461 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc 5521 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg 5581 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag 5641 gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat aaagcaatag 5701 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa 5761 actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg ccaacgcgcg 5821 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc 5881 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc 5941 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg 6001 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat 6061 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag 6121 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga 6181 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg 6241 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt 6301 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac 6361 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc 6421 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt 6481 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc 6541 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc 6601 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg 6661 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag 6721 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg 6781 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt 6841 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca 6901 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca 6961 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc 7021 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt 7081 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg 7141 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc 7201 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg 7261 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga 7321 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga 7381 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta 7441 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg 7501 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact 7561 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata 7621 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatgacc gctttggaaa 7681 aacaaagact gtatttcctg gaaattaatg tttattcaat aaactgtg //