LOCUS       sequence-209475-        5525 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of E. coli LF82_153 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5525)
  AUTHORS   Collins A, Huett A
  TITLE     A multi-phenotypic imaging screen to identify bacterial effectors by
            exogenous expression in a HeLa cell line.
  JOURNAL   Sci Data. 2018 May 15;5:180081. doi: 10.1038/sdata.2018.81.
  PUBMED    29762554
REFERENCE   2  (bases 1 to 5525)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Data.";
            date: "2018-05-15"; volume: "5:180081. doi"; pages: " 
            10.1038/sdata.2018.81"
COMMENT     Sequence Label: pCMV_mEm_4GS_LF82_153
FEATURES             Location/Qualifiers
     source          1..5525
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        44..347
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        348..551
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     502..522
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     548..572
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          689..785
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             join(846..848,849..851,852..1562)
                     /codon_start=1
                     /product="brightly fluorescent monomeric variant of GFP"
                     /label=mEmerald
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(891..912)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1152..1171)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1499..1520
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2681..2815
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2705..2724)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2759..2778
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2926..2942)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2926..2942)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2939..2961)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2950..2966
                     /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(join(2974..2980,2981..2998,2999..3004))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3019..3040
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3157..3174)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3328..3916)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3408..3427)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4087..4878,4879..4947))
                     /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     4710..4729
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4948..5052)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5120..5138
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5176..5198)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5298..5317
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(5511..5525,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(5525,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgag ttcgagcttg catgcctgca ggtcgttaca taacttacgg
       61 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      121 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      181 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      241 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
      301 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
      361 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
      421 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
      481 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
      541 taagcagagc tcgtttagtg aaccgtcaga tcgcctggag acgccatcca cgctgttttg
      601 acctccatag aagacaccgg gaccgatcca gcctccggac tctagaggat ccggtactag
      661 aggaactgaa aaaccagaaa gttaactggt aagtttagtc tttttgtctt ttatttcagg
      721 tcccggatcc ggtggtggtg caaatcaaag aactgctcct cagtggatgt tgcctttact
      781 tctaggcctg tacggaagtg ttacttctgc tctaaaagct gcggaattgt acccgcgggc
      841 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
      901 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
      961 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1021 ccaccctcgt gaccaccttg acctacggcg tgcagtgctt cgcccgctac cccgaccaca
     1081 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1141 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1201 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1261 ggcacaagct ggagtacaac tacaacagcc acaaggtcta tatcaccgcc gacaagcaga
     1321 agaacggcat caaggtgaac ttcaagaccc gccacaacat cgaggacggc agcgtgcagc
     1381 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     1441 accactacct gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca
     1501 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     1561 agcgaattgg tggagggggc ggttccggtg gcggaggttc ccgaattcgt cgacggagat
     1621 ctgatatgaa agataacaga caaaaagtcg ttcattataa aaaagcagta atcccacaca
     1681 gcaaatcatc attacaaaac atactactta gtattatagg tgagggcggc gttgcagaaa
     1741 aagtacttag tagacaagaa aaaataacac cttcagatga aaattcgggg tatcgtttct
     1801 taaataaaag cgacacatac aaaactatac tttttgggca actggtttta tttgaaaaag
     1861 acaaaagcca gtctctatta gagcttagtg ataatgttaa attttatgac attaactcca
     1921 tcacgtctaa tgatataatt attgatggag ataaacgtag tgacaaaaaa agagagtttg
     1981 tagattcaat tttatatttt ggtgtactaa acaaccatgt catgatcgtt caatccacat
     2041 cgcttcgagc aagggagctt gaggcgcatt tgaattggct aattaataca ttcacagatg
     2101 aaaaagaatc tgtgttaatg cttcaagata agccatccga agaaacgatg aaaaaaatgg
     2161 agaaagcacc agccaagagc ataaaaattg gtggattacc cattacttca gaagttaaaa
     2221 attcagagaa ccaaacaccc tccttttcaa atggttcggt gaagacaata aaataccgcc
     2281 caatagggaa aggaggtgcg ctactaaaag cattttttgg cgataactgg gcaaatgata
     2341 taaacattaa agattcatta gatgaggcca atttacaagt aagcctcgag attacatact
     2401 tcagaaaaac aaatgatagc ggtcaagctg ttatagacag tattgctacc tcactacgac
     2461 atctagatga ggatgatatt agcatcaaac ttcaaggcgg tggggaaata acgggaaaag
     2521 agataaaatt atccggaaaa attagcgttg aatataacaa tggcattatt gatgaaaatg
     2581 atttattctt aaaaatgcat aagtggctgt tctcaaaact tgatgccgga gaattggttg
     2641 caaaatgaat cggtaccctc gagaagcttg cggccgcggg gatccagaca tgataagata
     2701 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     2761 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     2821 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttcgga
     2881 tcctctagag tcgatctgca ggcatgctag cttggcgtaa tcatggtcat agctgtttcc
     2941 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     3001 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     3061 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     3121 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     3181 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     3241 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     3301 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     3361 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     3421 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     3481 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     3541 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     3601 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     3661 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     3721 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     3781 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3841 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     3901 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     3961 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     4021 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     4081 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     4141 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     4201 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     4261 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     4321 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     4381 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     4441 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     4501 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     4561 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     4621 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     4681 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     4741 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     4801 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     4861 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     4921 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     4981 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     5041 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     5101 catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg
     5161 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta
     5221 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
     5281 gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc atatgcggtg
     5341 tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgccatt cgccattcag
     5401 gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc
     5461 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg
     5521 acgtt
//
