LOCUS       sequence-209476-        5546 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of E. coli LF82_159 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5546)
  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 5546)
  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_159
FEATURES             Location/Qualifiers
     source          1..5546
                     /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"
     CDS             2143..2154
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     polyA_signal    2702..2836
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2726..2745)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2780..2799
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2947..2963)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2947..2963)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2960..2982)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2971..2987
                     /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(2995..3001,3002..3019,3020..3025))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3040..3061
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3178..3195)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3349..3937)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3429..3448)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4108..4899,4900..4968))
                     /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     4731..4750
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4969..5073)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5141..5159
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5197..5219)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5319..5338
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(5532..5546,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(5546,1..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 aattaacaag atattatcat ctgcaacact attgtatggt atgtcaatgg
     1681 ccatgtcggt cgggagttgt gcaacacctg tccagactaa tcttcctggt tacaccccgg
     1741 gtgcagatat cattagtgtt tcaccgacca gaaaccaggt cgatctcatt ggtgatgttg
     1801 tttattccca gataaaagga actcgttctg tcagacagct tcacatgtca gttcttgtcc
     1861 cgcgaacaaa tgatttaaaa ccagccatta tttattatcc cggcggcgga ttcatgtctt
     1921 ctgaacatga caaatttatt gaaatgagaa tggctctggc agaagctggt tttgttgtgg
     1981 ccgctgtaga atacagaaca attcctgata catttccagc accagttgag gatggaaaag
     2041 ctgcaatacg ttacctgaga gagcatgcca gcaattatgg gattgatcct caaagaatcg
     2101 gagttctggg tgactctgcc ggtggatggc ttgcccagat gatgggaact acaaatggtg
     2161 acaaaacctt tgataaaggt gactttcttc agcaatcctc agatgttcag gcagttgcca
     2221 cactttatgg gatttctgac ttgttgaata ttggcgaggg gttccctgaa tcagtgcagg
     2281 aggttcatcg ctctcctgcc gtaaccgaag ccttaatgat caatggccct gcattcagaa
     2341 attttgcggg agcccccatc acagcgtcaa aagaaaaagc gctaaacgcc agtccaatcg
     2401 gacatatgaa aggagtaaaa cccccatttc ttattatgca tggtagcaaa gacactctgg
     2461 tttcacctga gcaaagcgcc aaactattca ggatgttgaa gaagaacggt gataacgctg
     2521 agtacgtgct ggtagaaggg gccgagcatg gcgataatac atggtatcag ccaattatta
     2581 taaacagagt tgttgagtgg tttactaaaa acctgggaac ccccataaaa accgctaccc
     2641 aacaacaaaa aacaaacgct gacctgtgaa tcggtaccct cgagaagctt gcggccgcgg
     2701 ggatccagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg
     2761 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     2821 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     2881 ggtgtgggag gttttttcgg atcctctaga gtcgatctgc aggcatgcta gcttggcgta
     2941 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     3001 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     3061 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     3121 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     3181 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3241 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3301 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3361 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3421 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3481 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3541 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3601 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3661 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3721 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3781 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3841 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3901 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3961 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     4021 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     4081 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     4141 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     4201 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     4261 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     4321 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     4381 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     4441 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4501 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4561 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4621 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     4681 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     4741 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     4801 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     4861 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     4921 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     4981 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     5041 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     5101 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     5161 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     5221 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     5281 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     5341 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     5401 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     5461 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     5521 gccagggttt tcccagtcac gacgtt
//
