LOCUS Exported 7505 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Monomeric TagBFP2 and iCre-ERT2 recombinase (PAGSAS N-terminus). ACCESSION . VERSION . KEYWORDS pCAGGS-mTagBFP2-T2A-iCreERT2 (PAGSAS) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7505) AUTHORS Nam HS, Capecchi MR TITLE 2A creER optimization plasmids JOURNAL Unpublished REFERENCE 2 (bases 1 to 7505) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7505 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 11..390 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 392..669 /label=chicken beta-actin promoter intron 670..1686 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 1610..1632 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 1694..1713 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" CDS 1744..2454 /codon_start=1 /product="enhanced monomeric blue fluorescent protein (Subach et al., 2011)" /label=mTagBFP2 /note="mammalian codon-optimized" /translation="MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTM RIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG VLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRND MALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAV ARYCDLPSKLGHKLN" CDS 2476..2529 /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" CDS 3580..4515 /codon_start=1 /product="mutated ligand-binding domain of the human estrogen receptor (Feil et al., 1997)" /label=ERT2 /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEET DQSHLATAGSTSSHSLQKYYITGEAEGFPAT" primer_bind complement(4533..4552) /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" polyA_signal 4598..4653 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(4652..4671) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(5014..5030) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5014..5030) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5027..5049) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5038..5054 /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(5062..5092) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5107..5128 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5187..5382 /label=SV40 promoter /note="SV40 early promoter" rep_origin 5233..5368 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5295..5314 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 5388..5522 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5425..5444) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5479..5498 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5590..5607) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5761..6349) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5841..5860) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6520..7380) /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 7143..7162 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7381..7485) /gene="bla" /label=AmpR promoter ORIGIN 1 cacctgggtc gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta 61 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc 121 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg 181 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg 241 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa 301 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac 361 atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact 421 ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt 481 tgtgcagcga tgggggcggg gggggggggg gggcgcgcgc cggggggggg gggggggggg 541 gggggggggg gsggggsgrg gcggagaggt gcggcggcag ccaatcagag cggcgcgctc 601 cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg 661 cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg 721 cgccgcccgc cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc 781 ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg 841 ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg 901 ggtgcgtgcg tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc 961 tgtgagcgct gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc 1021 gcggccgggg gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg 1081 cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc 1141 cccctgcacc cccctccccg agttgctgag cacggcccgg cttcgggtgc ggggctccgt 1201 acggggcgtg gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg 1261 ggcggggcgg ggccgccycg ggccggggrg ggctcggggg rggggcgcgg cggcccccgg 1321 agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc 1381 gagagggcgc agggacttcc tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc 1441 cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg 1501 ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg 1561 ggctgtccgc ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc 1621 tggcgtgtga ccggcggctc tagagcctct gctaaccatg ttcatgcctt cttctttttc 1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattc 1741 gccatggtgt ctaagggcga agagctgatt aaggagaaca tgcacatgaa gctgtacatg 1801 gagggcaccg tggacaacca tcacttcaag tgcacatccg agggcgaagg caagccctac 1861 gagggcaccc agaccatgag aatcaaggtg gtcgagggcg gccctctccc cttcgccttc 1921 gacatcctgg ctactagctt cctctacggc agcaagacct tcatcaacca cacccagggc 1981 atccccgact tcttcaagca gtccttccct gagggcttca catgggagag agtcaccaca 2041 tacgaagacg ggggcgtgct gaccgctacc caggacacca gcctccagga cggctgcctc 2101 atctacaacg tcaagatcag aggggtgaac ttcacatcca acggccctgt gatgcagaag 2161 aaaacactcg gctgggaggc cttcaccgag acgctgtacc ccgctgacgg cggcctggaa 2221 ggcagaaacg acatggccct gaagctcgtg ggcgggagcc atctgatcgc aaacgccaag 2281 accacatata gatccaagaa acccgctaag aacctcaaga tgcctggcgt ctactatgtg 2341 gactacagac tggaaagaat caaggaggcc aacaacgaga cctacgtcga gcagcacgag 2401 gtggcagtgg ccagatactg cgacctccct agcaaactgg ggcacaagct taatggcggc 2461 tccggcggcc gctccgaggg ccgcggcagc ctgctgacat gcggcgacgt ggaagagaac 2521 cctggccctg ccggcagcgc cagcaacctg ctgactgtgc accaaaacct gcctgccctc 2581 cctgtggatg ccacctctga tgaagtcagg aagaacctga tggacatgtt cagggacagg 2641 caggccttct ctgaacacac ctggaagatg ctcctgtctg tgtgcagatc ctgggctgcc 2701 tggtgcaagc tgaacaacag gaaatggttc cctgctgaac ctgaggatgt gagggactac 2761 ctcctgtacc tgcaagccag aggcctggct gtgaagacca tccaacagca cctgggccag 2821 ctcaacatgc tgcacaggag atctggcctg cctcgccctt ctgactccaa tgctgtgtcc 2881 ctggtgatga ggagaatcag aaaggagaat gtggatgctg gggagagagc caagcaggcc 2941 ctggcctttg aacgcactga ctttgaccaa gtcagatccc tgatggagaa ctctgacaga 3001 tgccaggaca tcaggaacct ggccttcctg ggcattgcct acaacaccct gctgcgcatt 3061 gccgaaattg ccagaatcag agtgaaggac atctcccgca ccgatggtgg gagaatgctg 3121 atccacattg gcaggaccaa gaccctggtg tccacagctg gtgtggagaa ggccctgtcc 3181 ctgggggtta ccaagctggt ggagagatgg atctctgtgt ctggtgtggc tgatgacccc 3241 aacaactacc tgttctgccg ggtcagaaag aatggtgtgg ctgccccttc tgccacctcc 3301 caactgtcca cccgggccct ggaagggatc tttgaggcca cccaccgcct gatctatggt 3361 gccaaggatg actctgggca gagatacctg gcctggtctg gccactctgc cagagtgggt 3421 gctgccaggg acatggccag ggctggtgtg tccatccctg aaatcatgca ggctggtggc 3481 tggaccaatg tgaacattgt gatgaactac atcagaaacc tggactctga gactggggcc 3541 atggtgaggc tgctagagga tggggacctc gagccatctg ctggagacat gagagctgcc 3601 aacctttggc caagcccgct catgatcaaa cgctctaaga agaacagcct ggccttgtcc 3661 ctgacggccg accagatggt cagtgccttg ttggatgctg agccccccat actctattcc 3721 gagtatgatc ctaccagacc cttcagtgaa gcttcgatga tgggcttact gaccaacctg 3781 gcagacaggg agctggttca catgatcaac tgggcgaaga gggtgccagg ctttgtggat 3841 ttgaccctcc atgatcaggt ccaccttcta gaatgtgcct ggctagagat cctgatgatt 3901 ggtctcgtct ggcgctccat ggagcaccca gtgaagctac tgtttgctcc taacttgctc 3961 ttggacagga accagggaaa atgtgtagag ggcatggtgg agatcttcga catgctgctg 4021 gctacatcat ctcggttccg catgatgaat ctgcagggag aggagtttgt gtgcctcaaa 4081 tctattattt tgcttaattc tggagtgtac acatttctgt ccagcaccct gaagtctctg 4141 gaagagaagg accatatcca ccgagtcctg gacaagatca cagacacttt gatccacctg 4201 atggccaagg caggcctgac cctgcagcag cagcaccagc ggctggccca gctcctcctc 4261 atcctctccc acatcaggca catgagtaac aaaggcatgg agcatctgta cagcatgaag 4321 tgcaagaacg tggtgcccct ctatgacctg ctgctggagg cggcggacgc ccaccgccta 4381 catgcgccca ctagccgtgg aggggcatcc gtggaggaga cggaccaaag ccacttggcc 4441 actgcgggct ctacttcatc gcattccttg caaaagtatt acatcacggg ggaggcagag 4501 ggtttccctg ccacagcttg aggcgcgccg gatctttttc cctctgccaa aaattatggg 4561 gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt 4621 gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat 4681 catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg 4741 gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct 4801 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt 4861 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga 4921 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc 4981 ctcgacctgc agcccaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg 5041 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg 5101 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc 5161 gggaaacctg tcgtgccagc ggatccgcat ctcaattagt cagcaaccat agtcccgccc 5221 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc 5281 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag 5341 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctaac ttgtttattg 5401 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt 5461 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga 5521 tccgctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc 5581 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc 5641 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa 5701 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt 5761 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg 5821 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg 5881 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag 5941 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc 6001 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa 6061 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg 6121 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc 6181 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac 6241 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg 6301 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt 6361 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt 6421 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa 6481 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga 6541 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt 6601 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg 6661 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga 6721 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga 6781 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg 6841 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc 6901 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc 6961 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca 7021 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac 7081 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg 7141 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc 7201 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg 7261 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac 7321 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat 7381 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata 7441 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa 7501 agtgc //