LOCUS Exported 7356 bp ds-DNA circular SYN 25-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS CKAR SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7356) AUTHORS Violin JD, Zhang J, Tsien RY, Newton AC TITLE A genetically encoded fluorescent reporter reveals oscillatory phosphorylation by protein kinase C. JOURNAL J Cell Biol. 2003 Jun 9. 161(5):899-909. PUBMED 12782683 REFERENCE 2 (bases 1 to 7356) AUTHORS . TITLE Direct Submission JOURNAL Exported May 25, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7356 /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" regulatory 662..671 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 668..1384 /codon_start=1 /product="monomeric super enhanced variant of CFP (Matsuda et al., 2008)" /label=mseCFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" CDS 668..1384 /codon_start=1 /product="cyan variant of GFP" /label=CFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" CDS 668..1384 /codon_start=1 /product="enhanced CFP with monomerizing A206K mutation (Zacharias et al., 2002)" /label=mECFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" CDS 1943..2659 /codon_start=1 /product="enhanced YFP with monomerizing A206K mutation (Zacharias et al., 2002)" /label=mEYFP /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" promoter complement(2675..2693) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(2676..2693) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(2713..2730) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 2719..2943 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 2989..3417 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3076..3095) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3286..3307 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3426..3446) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3431..3760 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3611..3746 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3673..3692 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3827..4621 /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(3881..3900) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4491..4510 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 4795..4916 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4832..4851) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4886..4905 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(4965..4981) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4965..4981) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4978..5000) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4989..5005 /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(5013..5043) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5058..5079 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5196..5213) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5367..5955) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5447..5466) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6126..6986) /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 6749..6768 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6987..7091) /gene="bla" /label=AmpR promoter primer_bind complement(7166..7185) /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 acccaagctt 661 cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga 721 gctggacggc gacgtaaacg gccacaggtt cagcgtgtcc ggcgagggcg agggcgatgc 781 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg 841 gcccaccctc gtgaccaccc tgacctgggg cgtgcagtgc ttcagccgct accccgacca 901 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgtac 961 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga 1021 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct 1081 ggggcacaag ctggagtaca actacatcag ccacaacgtc tatatcaccg ccgacaagca 1141 gaagaacggc atcaaggccc acttcaagat ccgccacaac atcgaggacg gcagcgtgca 1201 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga 1261 caaccactac ctgagcaccc agtccaagct gagcaaagac cccaacgaga agcgcgatca 1321 catggtcctg ctggagttcg tgaccgccgc cgggatcact cacggcatgg acgagctgta 1381 caaggctggt accaagggta atggtaggtt tttaacttta aaaccattgc ctgacagcat 1441 tattcaagaa agcctggaga ttcagcaagg tgtgaatcca tttttcattg gtagatccga 1501 ggattgcaat tgtaaaattg aagacagtag gttgtctcga gttcattgct tcattttcaa 1561 aaagaggcat gctgtaggca aaagcatgta tgaatctccg gcacaaggtt tagatgatat 1621 ttggtattgc cacaccggaa ctaacgtgag ctatttaaat aataaccgca tgatacaggg 1681 tacgaaattc cttttacaag acggagatga aatcaagatc atttgggata aaaacaataa 1741 atttgtcatt ggctttaaag tggaaattaa cgatactaca ggtctgttta acgagggatt 1801 aggtatgtta caagaacaaa gagtagtact taagcaaaca gccgaagaaa aagatttggt 1861 gaaaaagtta ggaggatccg gaggtcgatt cagaagattc cagacgttga agatcaaggc 1921 taaggcaggt ggatctggtg gagtgagcaa gggcgaggag ctgttcaccg gggtggtgcc 1981 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg 2041 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct 2101 gcccgtgccc tggcccaccc tcgtgaccac cttcggctac ggcctgatgt gcttcgcccg 2161 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt 2221 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa 2281 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga 2341 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat 2401 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga 2461 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt 2521 gctgctgccc gacaaccact acctgagcta ccagtccaag ctgagcaaag accccaacga 2581 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctcacggcat 2641 ggacgagctg tacaagtaat ctagagggcc ctattctata gtgtcaccta aatgctagag 2701 ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc 2761 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg 2821 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg 2881 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta 2941 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta 3001 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca 3061 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct 3121 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc 3181 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat 3241 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc 3301 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc 3361 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat 3421 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag 3481 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc 3541 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct 3601 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg 3661 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa 3721 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat 3781 atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga 3841 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc 3901 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc 3961 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc 4021 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac 4081 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc 4141 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac 4201 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg 4261 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct 4321 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt 4381 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg 4441 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac 4501 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg 4561 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg 4621 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat 4681 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc 4741 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg 4801 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa 4861 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat 4921 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct 4981 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt 5041 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc 5101 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg 5161 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg 5221 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca 5281 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac 5341 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac 5401 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg 5461 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac 5521 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat 5581 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag 5641 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac 5701 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt 5761 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt 5821 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc 5881 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga 5941 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac 6001 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc 6061 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct 6121 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca 6181 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct 6241 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca 6301 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc 6361 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg 6421 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct 6481 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa 6541 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta 6601 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc 6661 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg 6721 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa 6781 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg 6841 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc 6901 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg 6961 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat 7021 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata 7081 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc 7141 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt 7201 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta 7261 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg 7321 cgctgcttcg cgatgtacgg gccagatata cgcgtt //