LOCUS Exported 6558 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS 223 pCS EGFP DEST SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6558) AUTHORS Villefranc JA, Amigo J, Lawson ND TITLE Gateway compatible vectors for analysis of gene function in the zebrafish. JOURNAL Dev Dyn. 2007 Nov . 236(11):3077-87. PUBMED 17948311 REFERENCE 2 (bases 1 to 6558) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6558 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 35..53 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 35..52 /label=SP6 /note="SP6 promoter, forward primer" regulatory 119..128 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 125..841 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 125..841 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(170..191) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(431..450) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 778..799 /label=EGFP-C /note="EGFP, forward primer" protein_bind 863..987 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 1012..1042 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 1096..1755 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(1162..1181) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 2097..2402 /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind 2281..2300 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(2443..2567) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter complement(2585..2602) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" polyA_signal 2607..2741 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2631..2650) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2685..2704 /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind complement(2854..2874) /label=T3 /note="T3 promoter, forward primer" promoter complement(2854..2872) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(2893..2909) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2893..2909) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(2906..2928) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 2917..2933 /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(2941..2971) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 2986..3007 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(3124..3141) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3295..3883) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3375..3394) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4054..4914) /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 4677..4696 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(4915..5019) /gene="bla" /label=AmpR promoter rep_origin complement(5045..5500) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5182..5203) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5394..5413 /label=F1ori-R /note="F1 origin, reverse primer" promoter 5574..6558 /label=CMV IE94 promoter /note="enhancer/promoter region of simian cytomegalovirus major immediate early transcription unit IE94" ORIGIN 1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag 61 ctacttgttc tttttgcagg atcccatcga ttcgaattca aggctagcgc taccggtcgc 121 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct 181 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac 241 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc 301 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat 361 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat 421 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac 481 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg 541 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa 601 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct 661 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa 721 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat 781 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa 841 gtccggactc agatctcgaa tcacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa 901 tgatataaat atcaatatat taaattagat tttgcataaa aaacagacta cataatactg 961 taaaacacaa catatccagt cactatggcg gccgcattag gcaccccagg ctttacactt 1021 tatgcttccg gctcgtataa tgtgtggatt ttgagttagg atccgtcgag attttcagga 1081 gctaaggaag ctaaaatgga gaaaaaaatc actggatata ccaccgttga tatatcccaa 1141 tggcatcgta aagaacattt tgaggcattt cagtcagttg ctcaatgtac ctataaccag 1201 accgttcagc tggatattac ggccttttta aagaccgtaa agaaaaataa gcacaagttt 1261 tatccggcct ttattcacat tcttgcccgc ctgatgaatg ctcatccgga attccgtatg 1321 gcaatgaaag acggtgagct ggtgatatgg gatagtgttc acccttgtta caccgttttc 1381 catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga tttccggcag 1441 tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc ctatttccct 1501 aaagggttta ttgagaatat gtttttcgtc tcagccaatc cctgggtgag tttcaccagt 1561 tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac catgggcaaa 1621 tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtt 1681 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg 1741 cagggcgggg cgtaaacgcg tggatccggc ttactaaaag ccagataaca gtatgcgtat 1801 ttgcgcgctg atttttgcgg tataagaata tatactgata tgtatacccg aagtatgtca 1861 aaaagaggta tgctatgaag cagcgtatta cagtgacagt tgacagcgac agctatcagt 1921 tgctcaaggc atatatgatg tcaatatctc cggtctggta agcacaacca tgcagaatga 1981 agcccgtcgt ctgcgtgccg aacgctggaa agcggaaaat caggaaggga tggctgaggt 2041 cgcccggttt attgaaatga acggctcttt tgctgacgag aacaggggct ggtgaaatgc 2101 agtttaaggt ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga 2161 gtgatattat tgacacgccc gggcgacgga tggtgatccc cctggccagt gcacgtctgc 2221 tgtcagataa agtctcccgt gaactttacc cggtggtgca tatcggggat gaaagctggc 2281 gcatgatgac caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg 2341 atctcagcca ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat 2401 aaatgtcagg ctcccttata cacagccagt ctgcaggtcg accatagtga ctggatatgt 2461 tgtgttttac agcattatgt agtctgtttt ttatgcaaaa tctaatttaa tatattgata 2521 tttatatcat tttacgtttc tcgttcagct ttcttgtaca aagtggtgat tcgagcctct 2581 agaactatag tgagtcgtat tacgtagatc cagacatgat aagatacatt gatgagtttg 2641 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta 2701 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc 2761 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaattcgcg gccgcggcgc 2821 caatgcattg ggcccggtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt 2881 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca 2941 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact 3001 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct 3061 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc 3121 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca 3181 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg 3241 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca 3301 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa 3361 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc 3421 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc 3481 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct 3541 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg 3601 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag 3661 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta 3721 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg 3781 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt 3841 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt 3901 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag 3961 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat 4021 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc 4081 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat 4141 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc 4201 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag 4261 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag 4321 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt 4381 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg 4441 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt 4501 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc 4561 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc 4621 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa 4681 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg 4741 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc 4801 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag 4861 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt 4921 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt 4981 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc 5041 acctaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc 5101 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc 5161 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac 5221 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca 5281 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg 5341 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag 5401 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc 5461 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg 5521 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gtcgaccata 5581 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt 5641 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca 5701 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ttggcactgt 5761 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg 5821 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat 5881 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca 5941 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt 6001 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac 6061 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac 6121 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta 6181 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa 6241 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg 6301 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac 6361 tattggaagg acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg 6421 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa 6481 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc 6541 aatgctcgtt tagggaac //