LOCUS       sequence-209650-        4691 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of E. coli LF82_147 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4691)
  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 4691)
  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_147
FEATURES             Location/Qualifiers
     source          1..4691
                     /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    1847..1981
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1871..1890)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1925..1944
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2092..2108)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2092..2108)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2105..2127)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2116..2132
                     /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(2140..2146,2147..2164,2165..2170))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2185..2206
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2323..2340)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2494..3082)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2574..2593)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3253..4044,4045..4113))
                     /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     3876..3895
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4114..4218)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4286..4304
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4342..4364)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4464..4483
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(4677..4691,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(4691,1..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 ctgatatggc gatgacgcat cctcacgata atatccgggt aggcgcgatc actttcgtct
     1681 actccgttac aaagcgaggc tgggtatttc ccggcctttc tgttatccaa aatccactga
     1741 aagcccagcg gctggctgag gagataaata ataaacgggg ggctgtatgc acaaagcatc
     1801 tcccgttgag ttgaatcggt accctcgaga agcttgcggc cgcggggatc cagacatgat
     1861 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     1921 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     1981 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt
     2041 ttcggatcct ctagagtcga tctgcaggca tgctagcttg gcgtaatcat ggtcatagct
     2101 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     2161 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     2221 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     2281 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     2341 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     2401 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     2461 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     2521 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2581 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2641 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2701 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2761 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2821 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2881 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     2941 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3001 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3061 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3121 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3181 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3241 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3301 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3361 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     3421 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3481 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3541 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3601 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3661 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3721 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3781 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     3841 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3901 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3961 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4021 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4081 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4141 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4201 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     4261 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     4321 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     4381 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     4441 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     4501 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc
     4561 attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca
     4621 gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca
     4681 gtcacgacgt t
//
