LOCUS Exported 6127 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION GFP with robust performance in protein fusions and useful as FRET acceptor for mRuby2 in a FRET pair that offers bright fluorescence, dynamic range, and photostability while limiting emissions overlap. ACCESSION . VERSION . KEYWORDS pcDNA3-Clover SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6127) AUTHORS Lam AJ, St-Pierre F, Gong Y, Marshall JD, Cranfill PJ, Baird MA, McKeown MR, Wiedenmann J, Davidson MW, Schnitzer MJ, Tsien RY, Lin MZ TITLE Improving FRET dynamic range with bright green and red fluorescent proteins. JOURNAL Nat Methods. 2012 Sep 9. doi: 10.1038/nmeth.2171. PUBMED 22961245 REFERENCE 2 (bases 1 to 6127) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6127 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(44..63) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 235..614 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 615..818 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 769..789 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 863..882 /label=T7 /note="T7 promoter, forward primer" promoter 863..881 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 947..1663 /codon_start=1 /product="bright green-yellow fluorescent protein derived from GFP (Lam et al., 2012)" /label=Clover /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" primer_bind complement(992..1013) /label=EGFP-N /note="EGFP, reverse primer" primer_bind 1600..1621 /label=EGFP-C /note="EGFP, forward primer" promoter complement(1679..1697) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(1680..1697) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(1717..1734) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 1723..1947 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 1993..2421 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2080..2099) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2290..2311 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(2430..2450) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 2435..2765 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 2616..2751 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2678..2697 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 2832..3626 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(2886..2905) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3496..3515 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 3800..3921 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3837..3856) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 3891..3910 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(3970..3986) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3970..3986) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(3983..4005) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 3994..4010 /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(4018..4048) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4063..4084 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4201..4218) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4372..4960) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4452..4471) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5131..5991) /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 5754..5773 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5992..6096) /gene="bla" /label=AmpR promoter ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc 901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcggcatgg tgagcaaggg 961 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg 1021 ccacaagttc agcgtccgcg gcgagggcga gggcgatgcc accaacggca agctgaccct 1081 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt 1141 cggctacggc gtggcctgct tcagccgcta ccccgaccac atgaagcagc acgacttctt 1201 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atctctttca aggacgacgg 1261 tacctacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga 1321 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa 1381 cttcaacagc cacaacgtct atatcacggc cgacaagcag aagaacggca tcaaggctaa 1441 cttcaagatc cgccacaacg ttgaggacgg cagcgtgcag ctcgccgacc actaccagca 1501 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagccatca 1561 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt 1621 gaccgccgcc gggattacac atggcatgga cgagctgtac aagtctagag ggccctattc 1681 tatagtgtca cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc 1741 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca 1801 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta 1861 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc 1921 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta 1981 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc 2041 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt 2101 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggc atccctttag 2161 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt 2221 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt 2281 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt 2341 cttttgattt ataagggatt ttggggattt cggcctattg gttaaaaaat gagctgattt 2401 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc 2461 cccaggctcc ccaggcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca 2521 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt 2581 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt 2641 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg 2701 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt 2761 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag 2821 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg 2881 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt 2941 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc 3001 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt 3061 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag 3121 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg 3181 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag 3241 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg 3301 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc 3361 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca 3421 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc 3481 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg 3541 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct 3601 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc 3661 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg 3721 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct 3781 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa 3841 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc 3901 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc 3961 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa 4021 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac 4081 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca 4141 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc 4201 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc 4261 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc 4321 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag 4381 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc 4441 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt 4501 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct 4561 ttctcaatgc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg 4621 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct 4681 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat 4741 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg 4801 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa 4861 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt 4921 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc 4981 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt 5041 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta 5101 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat 5161 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac 5221 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg 5281 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag 5341 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt 5401 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt 5461 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt 5521 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt 5581 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct 5641 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt 5701 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac 5761 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa 5821 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa 5881 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca 5941 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct 6001 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga 6061 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc 6121 tgacgtc //