LOCUS Exported 3630 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Constitutive expression of mClover3 in bacteria. ACCESSION . VERSION . KEYWORDS pNCS-mClover3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3630) AUTHORS Bajar BT, Wang ES, Lam AJ, Kim BB, Jacobs CL, Howe ES, Davidson MW, Lin MZ, Chu J TITLE Improving brightness and photostability of green and red fluorescent proteins for live cell imaging and FRET reporting. JOURNAL Sci Rep. 2016 Feb 16;6:20889. doi: 10.1038/srep20889. PUBMED 26879144 REFERENCE 2 (bases 1 to 3630) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3630 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 20..39 /label=T7 /note="T7 promoter, forward primer" promoter 20..38 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 164..181 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 185..217 /codon_start=1 /product="leader peptide from bacteriophage T7 gene 10" /label=T7 tag (gene 10 leader) /note="promotes efficient translation in E. coli" /translation="MASMTGGQQMG" CDS 221..244 /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" CDS 251..970 /codon_start=1 /product="bright green-yellow fluorescent protein derived from mClover2, with improved photostability (Bajar et al., 2003)" /label=mClover3 /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" primer_bind complement(296..317) /label=EGFP-N /note="EGFP, reverse primer" primer_bind 904..925 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(1031..1049) /label=T7 Term /note="T7 terminator, reverse primer" terminator 1045..1092 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 1189..1644 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1276..1295) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 1486..1507 /label=F1ori-F /note="F1 origin, forward primer" promoter 1670..1774 /gene="bla" /label=AmpR promoter CDS 1775..2635 /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(1993..2012) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 2806..3394 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3295..3314 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3548..3565 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct 61 agaatcaccg agtttattct tgacacctga tgcgatgaat gatataatag gaaagtactg 121 ttttgtttaa ctttaagaag gggatataca tatgcggggt tctcatcatc atcatcatca 181 tggtatggct agcatgactg gtggacagca aatgggtcgg gatctgtacg acgatgacga 241 taaggatccg atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt 301 cgagctggac ggcgacgtaa acggccacaa gttcagcgtc cgcggcgagg gcgagggcga 361 tgccaccaac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc 421 ctggcccacc ctcgtgacca ccttcggcta cggcgtggcc tgcttcagcc gctaccccga 481 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg 541 caccatctct ttcaaggacg acggtaccta caagacccgc gccgaggtga agttcgaggg 601 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat 661 cctggggcac aagctggagt acaacttcaa cagccactac gtctatatca cggccgacaa 721 gcagaagaac tgcatcaagg ctaacttcaa gatccgccac aacgttgagg acggcagcgt 781 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc 841 cgacaaccac tacctgagcc atcagtccaa gctgagcaaa gaccccaacg agaagcgcga 901 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatt acacatggca tggacgagct 961 gtacaagtga attcgaagct tgatccggct gctaacaaag cccgaaagga agctgagttg 1021 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg 1081 aggggttttt tgctgaaagg aggaactata tccggatctg gcgtaatagc gaagaggccc 1141 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta 1201 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca 1261 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct 1321 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc 1381 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat 1441 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc 1501 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc 1561 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta 1621 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc 1681 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg 1741 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 1801 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 1861 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct 1921 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 1981 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 2041 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 2101 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 2161 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 2221 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 2281 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 2341 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 2401 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 2461 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 2521 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 2581 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 2641 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 2701 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc 2761 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt 2821 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt 2881 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 2941 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 3001 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 3061 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 3121 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 3181 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 3241 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 3301 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 3361 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 3421 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 3481 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 3541 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 3601 ccccgcgcgt tggccgattc attaatgcag //