LOCUS Exported 3359 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Contributes the coding sequence for ECFP as the 3'-module during MultiSite Gateway-cloning of chimeric cDNAs encoding three-part fusion proteins.. ACCESSION . VERSION . KEYWORDS pDONR P2R-P3-ECFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3359) AUTHORS Buj R, Iglesias N, Planas AM, Santalucia T TITLE A plasmid toolkit for cloning chimeric cDNAs encoding customized fusion proteins into any Gateway destination expression vector. JOURNAL BMC Mol Biol. 2013 Aug 20;14(1):18. doi: 10.1186/1471-2199-14-18. PUBMED 23957834 REFERENCE 2 (bases 1 to 3359) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3359 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 92..109 /label=L4440 /note="L4440 vector, forward primer" terminator 268..295 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" terminator 387..473 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" primer_bind 522..544 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 536..553 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 537..553 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" protein_bind complement(624..748) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" CDS 750..1466 /codon_start=1 /product="enhanced CFP" /label=ECFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELY" primer_bind complement(795..816) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1056..1075) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1403..1424 /label=EGFP-C /note="EGFP, forward primer" protein_bind complement(1468..1563) /gene="mutant version of attL" /label=attL3 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" primer_bind complement(1600..1619) /label=T7 /note="T7 promoter, forward primer" promoter complement(1601..1619) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1624..1640) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1624..1640) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1742..1761) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 1753..2562 /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind complement(1820..1839) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" rep_origin 2709..3297 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3198..3217 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga 61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga 121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca 181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc 241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta 301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc 361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa 421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg 481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa 541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac 601 taataccatc taagtagttg gttcatagtg actgcatatg ttgtgtttta cagtattatg 661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt 721 ctcgttcaac tttcttgtac aaagtggaca tggtgagcaa gggcgaggag ctgttcaccg 781 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 841 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 901 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctgg ggcgtgcagt 961 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1021 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1081 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1141 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacatc agccacaacg 1201 tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca 1261 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1321 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag 1381 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1441 ctctcggcat ggacgagctg tactaaacaa ctttattata caaagttggc attataaaaa 1501 agcattgctt atcaatttgt tgcaacgaac aggtcactat cagtcaaaat aaaatcatta 1561 tttggagctc catggtagcg ttaacgcggc cgcgatatcc cctatagtga gtcgtattac 1621 atggtcatag ctgtttcctg gcagctctgg cccgtgtctc aaaatctctg atgttacatt 1681 gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat aaacagtaat 1741 acaaggggtg ttatgagcca tattcaacgg gaaacgtcga ggccgcgatt aaattccaac 1801 atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg 1861 acaatctatc gcttgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa 1921 ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt 1981 atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc 2041 actgcgatcc ccggaaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa 2101 aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat 2161 tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac 2221 ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc 2281 tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat 2341 ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga 2401 cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag 2461 ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg 2521 aataaattgc agtttcattt gatgctcgat gagtttttct aatcagaatt ggttaattgg 2581 ttgtaacact ggcagagcat tacgctgact tgacgggacg gcgcaagctc atgaccaaaa 2641 tcccttaacg tgagttacgc gtcgttccac tgagcgtcag accccgtaga aaagatcaaa 2701 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca 2761 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta 2821 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc 2881 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca 2941 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta 3001 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag 3061 cgaacgacct acaccgaact gagataccta cagcgtgagc attgagaaag cgccacgctt 3121 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc 3181 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac 3241 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac 3301 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgtt //