LOCUS       sequence-210327-        5408 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of E. coli LF82_665 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5408)
  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 5408)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 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_665
FEATURES             Location/Qualifiers
     source          1..5408
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          126..222
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             join(283..285,286..288,289..999)
                     /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(328..349)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(589..608)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     936..957
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2001..2135
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2025..2044)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2079..2098
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2246..2262)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2246..2262)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2259..2281)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2270..2286
                     /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(2294..2300,2301..2318,2319..2324))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2339..2360
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2477..2494)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2648..3236)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2728..2747)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3407..4198,4199..4267))
                     /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     4030..4049
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4268..4372)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4440..4458
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4496..4518)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4618..4637
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4831..4853
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4845..4862
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4846..4862
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        4889..5192
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5193..5396
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5347..5367
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     join(5393..5408,1..9)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
ORIGIN
        1 cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac
       61 cgatccagcc tccggactct agaggatccg gtactagagg aactgaaaaa ccagaaagtt
      121 aactggtaag tttagtcttt ttgtctttta tttcaggtcc cggatccggt ggtggtgcaa
      181 atcaaagaac tgctcctcag tggatgttgc ctttacttct aggcctgtac ggaagtgtta
      241 cttctgctct aaaagctgcg gaattgtacc cgcgggccca ccatggtgag caagggcgag
      301 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
      361 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
      421 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccttgacc
      481 tacggcgtgc agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag
      541 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
      601 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
      661 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
      721 aacagccaca aggtctatat caccgccgac aagcagaaga acggcatcaa ggtgaacttc
      781 aagacccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
      841 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
      901 aagctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
      961 gccgccggga tcactctcgg catggacgag ctgtacaagc gaattggtgg agggggcggt
     1021 tccggtggcg gaggttcccg aattcgtcga cggagatctg atatgaatgt ctataaactc
     1081 tcagttcaga cccagggcta ttttcttatc gaaaagaacg attatgagca gagaactgcg
     1141 cgacgtcccc cctggtataa aatgagtgac gatggcgagc caagatactt cgccatttgc
     1201 ccggcctgca ataatccaac gcagattatt ggcttatata aactgcctgc gaatgtttat
     1261 cacccttatg ccaagcacta tggcaaacca attccgggag ttgggatata tgacagggaa
     1321 gcctatgagt ggtgtcccta ttcgaacagg aagccagcca gcaacgggaa taaaccaaag
     1381 cgtttgccgg gtgtactgac acaacagatc ctctccatac tggtgaccca ttttgacaaa
     1441 gtcgtctggt tattaagtca atccaccggt atcagtattg atgagaaact tgcccgaaag
     1501 atgcttgagc aatacaggcg ggaggacggg tggctttact cccgggcaac gttaatgaat
     1561 attccctggg cctttgccta catgacggat aaccaggaca tacttttcaa aaaattcggc
     1621 gatagcacac ttgcgaatga gcttgtaacc cgtgcaccgg agattctgga atttacggac
     1681 aagagatacc ttattaagca aaaaacctgt aaggagtata tcgagctggg tgcatattat
     1741 accttgcacc aatcaagggt ggttgatgat gttctgacag aatcaatgga catggttttt
     1801 accctaaaaa aaggatacag ggagccgcta gagatttatc gtcagactat ttcttttgat
     1861 tttgagcatt ttaataatct ggtcaatttc agcgagtgga aaacatcccc ctggaatgaa
     1921 caacttctcc agcttgcccg ggaaatcctg ggggatttga tcgcttgaat cggtaccctc
     1981 gagaagcttg cggccgcggg gatccagaca tgataagata cattgatgag tttggacaaa
     2041 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     2101 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     2161 tgtttcaggt tcagggggag gtgtgggagg ttttttcgga tcctctagag tcgatctgca
     2221 ggcatgctag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     2281 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     2341 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     2401 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     2461 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     2521 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     2581 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     2641 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     2701 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     2761 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     2821 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     2881 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     2941 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     3001 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     3061 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     3121 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     3181 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     3241 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     3301 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     3361 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     3421 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     3481 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     3541 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     3601 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     3661 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     3721 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     3781 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     3841 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     3901 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     3961 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     4021 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     4081 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     4141 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     4201 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     4261 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     4321 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     4381 cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
     4441 ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct
     4501 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     4561 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg
     4621 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     4681 taaggagaaa ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag
     4741 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     4801 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     4861 gtgagttcga gcttgcatgc ctgcaggtcg ttacataact tacggtaaat ggcccgcctg
     4921 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
     4981 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
     5041 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
     5101 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
     5161 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
     5221 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
     5281 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
     5341 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt
     5401 tagtgaac
//
