LOCUS Exported 6076 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses GFP(1-10) in mammalian cells. ACCESSION . VERSION . KEYWORDS pcDNA3.1-GFP(1-10) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6076) AUTHORS Kamiyama D, Sekine S, Barsi-Rhyne B, Hu J, Chen B, Gilbert LA, Ishikawa H, Leonetti MD, Marshall WF, Weissman JS, Huang B TITLE Versatile protein tagging in cells with split fluorescent protein. JOURNAL Nat Commun. 2016 Mar 18;7:11046. doi: 10.1038/ncomms11046. PUBMED 26988139 REFERENCE 2 (bases 1 to 6076) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6076 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..380 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 381..584 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 535..555 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 629..648 /label=T7 /note="T7 promoter, forward primer" promoter 629..647 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 736..1377 /codon_start=1 /product="fragment of superfolder GFP consisting of the first 10 beta-strands" /label=GFP(1-10) /note="associates with GFP11, the 11th beta-strand, to reconstitute fluorescent GFP (Cabantous et al., 2005; Kamiyama et al., 2016)" /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATIGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG KYKTRAVVKFEGDTLVNRIELKGTDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA NFTVRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQTVLSKDPNEK" primer_bind complement(1433..1450) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 1439..1663 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 1709..2137 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1796..1815) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2006..2027 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(2146..2166) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 2151..2480 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 2331..2466 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2393..2412 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 2547..3341 /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(2601..2620) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3211..3230 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 3515..3636 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3552..3571) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 3606..3625 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(3685..3701) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3685..3701) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(3698..3720) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 3709..3725 /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(3733..3763) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3778..3799 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(3916..3933) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4087..4675) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4167..4186) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4846..5706) /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 5469..5488 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5707..5811) /gene="bla" /label=AmpR promoter primer_bind complement(5886..5905) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag 601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg 661 gctagcgttt aaacttaagc ttggtaccga gctcggatcc actagtccag tgtggtggaa 721 ttcgttaaca gatctatgtc caaaggagaa gaactgttta ccggtgttgt gccaattttg 781 gttgaactcg atggtgatgt caacggacat aagttctcag tgagaggcga aggagaaggt 841 gacgccacca ttggaaaatt gactcttaaa ttcatctgta ctactggtaa acttcctgta 901 ccatggccga ctctcgtaac aacgcttacg tacggagttc agtgcttttc gagataccca 961 gaccatatga aaagacatga cttttttaag tcggctatgc ctgaaggtta cgtgcaagaa 1021 agaacaattt cgttcaaaga tgatggaaaa tataaaacta gagcagttgt taaatttgaa 1081 ggagatactt tggttaaccg cattgaactg aaaggaacag attttaaaga agatggtaat 1141 attcttggac acaaactcga atacaatttt aatagtcata acgtatacat cactgctgat 1201 aagcaaaaga acggaattaa agcgaatttc acagtacgcc ataatgtaga agatggcagt 1261 gttcaacttg ccgaccatta ccaacaaaac acccctattg gagacggtcc ggtacttctt 1321 cctgataatc actacctctc aacacaaaca gtcctgagca aagatccaaa tgaaaaagga 1381 acaggtggcg gcggaagtta gtctagaggg cccgtttaaa cccgctgatc agcctcgact 1441 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg 1501 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg 1561 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg 1621 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga 1681 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg 1741 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct 1801 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat 1861 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt 1921 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg 1981 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac 2041 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta 2101 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt 2161 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca 2221 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa 2281 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc 2341 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg 2401 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg 2461 gaggcctagg cttttgcaaa aagctcccgg gagcttatat atccattttc ggatctgatc 2521 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc 2581 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct 2641 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg 2701 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca 2761 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc 2821 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga 2881 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc 2941 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc 3001 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg 3061 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct 3121 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc 3181 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc 3241 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc 3301 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga 3361 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt 3421 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg 3481 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg 3541 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc 3601 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc 3661 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct 3721 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg 3781 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct 3841 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg 3901 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc 3961 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg 4021 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct 4081 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca 4141 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct 4201 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc 4261 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt 4321 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc 4381 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc 4441 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg 4501 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc 4561 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag 4621 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga 4681 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat 4741 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag 4801 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat 4861 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc 4921 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat 4981 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag 5041 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg 5101 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc 5161 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca 5221 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg 5281 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc 5341 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta 5401 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc 5461 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg 5521 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc 5581 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc 5641 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat 5701 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag 5761 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc 5821 ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact 5881 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg 5941 ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc 6001 gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg 6061 gccagatata cgcgtt //