LOCUS       sequence-212780-        8405 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of E. coli LF82_315 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8405)
  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 8405)
  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_315
FEATURES             Location/Qualifiers
     source          1..8405
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(96..115)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     443..464
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4505..4639
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4529..4548)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4583..4602
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4750..4766)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4750..4766)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4763..4785)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4774..4790
                     /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(4798..4804,4805..4822,4823..4828))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4843..4864
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4981..4998)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5152..5740)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5232..5251)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5911..6702,6703..6771))
                     /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     6534..6553
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6772..6876)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6944..6962
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7000..7022)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7122..7141
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7335..7357
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7349..7366
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7350..7366
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        7393..7696
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7697..7900
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7851..7871
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     7897..7921
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          8038..8134
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             join(8195..8197,8198..8200,8201..8405,1..506)
                     /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(8240..8261)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
       61 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
      121 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
      181 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaagg
      241 tctatatcac cgccgacaag cagaagaacg gcatcaaggt gaacttcaag acccgccaca
      301 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
      361 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag ctgagcaaag
      421 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
      481 ctctcggcat ggacgagctg tacaagcgaa ttggtggagg gggcggttcc ggtggcggag
      541 gttcccgaat tcgtcgacgg agatctgata tgagtgcccc tttaatacgt tcctacaatg
      601 ccgttattga atacaccctt tctcaggcaa aaaaatccaa aacatcccgt cgtatcgcag
      661 gattttgtca gcttcagggg cctacgggta gcggaaaaac cagctcactc taccgctctg
      721 gctatgccga taacgctctc ccggcgctgg agacgataaa aaaatcaggc tctcaagcca
      781 ttctggttac ccatcgctgg aatattttgc acgacattta tgagaaaacg gcagcgcata
      841 aagacagtaa cggggaacca ttcaccgtca gcgtgctcta tgcacaagat gaacagatcg
      901 tttctgccat cgaggccacc cccttgcctc atgagaataa tatttccgca gactcacttc
      961 ccgatccatt tgactccatc aatatcctgg aagacagaaa tttgtttgcc agtcaggata
     1021 tcgccgacaa attgcgtaga aattgcaaaa acatactgca tattaacaaa tcgctgaaag
     1081 gatatccgac gcgttttcag acggccatcg ccagtgagaa agagtcatta atcaaatcat
     1141 gcgcagcggt ggagataact ctgattcaga atatcaaatt gctggaaaaa gaagccaaga
     1201 aacaaaaaga aaaatatggc gaagaacatg aactcacgca ggcagccagg caacgagttg
     1261 cagatttccg tcgccacgtc tgggtcaggc gcattctgcc cgccattgcc tggcgtgacg
     1321 aaaagcaaca tctgctgatc atgacgactc aaaagatggt ttcatcgttt tacgatgggc
     1381 atcgcaaagt aaaaatgtcc agcaaagagc tgcgtaatta catcattttc atcgatgaat
     1441 ttgattacca gtctgaggta ttacaagagt atctggcaca ggcgcaatgg gttcaggaac
     1501 cacctgaatg tctgggtcag ttgctcgacg ggggacgcag attactacaa cgtatgcagt
     1561 atgttgaaac ggaacctgca cctatgatcc gtgaacgtct agaaaaattt attaatgatc
     1621 tggatagcgc attaacagat aaacatgttg atttaacaca tgccagatcg ctggttattc
     1681 ctcttcagca atatctggat aataaaccct ttggtgagca ttatttattc cgttccgatc
     1741 aactggtgac cagtgaacgg cttattatgc ggaaagcgga acatggctac gaaattatca
     1801 ggccagactc tccgcttcag gatagcgatc agactatcga catcagtgat tttttacgct
     1861 tgatggaaaa attcattcga cagtttagcc taatgttgac cgacctgact gcaagtgaag
     1921 atgaagcgta tgaatacatt aaaaagttaa cccgacttct gttcgacccc gttaatgatt
     1981 acaggccatc atactacggc tcaaccctgc ccaatatgtc tcgatttttg ttaccgcgta
     2041 ctgacctgcc ggaactacgg gaattaagga aaagtaatat tttgccaaat actcatgcca
     2101 gtatttttgg cttgaccaat tggttgctga aacagaatgc ctcagatacg gatatcgatc
     2161 cgttacggat acagatcaaa cgtgcctttc tacccacaac ggcagaaggt ctgctgcttt
     2221 ctctcgccag tcgtaatctt gtctttgcac tgtcggctac gtcatatatt gaaagagcct
     2281 gtaatcactt cgatgtccgg tggattaatt cagctcttcg ttatattgca gaagcacgca
     2341 accctgctgt tacacaaagt tttctgggaa caacttttga aaaacgacct gtcgagtggt
     2401 ttaaagaacc gataccttat gtccagacgg ctgatgattt ccagttgcag tatttagcta
     2461 ttgaagagat caataacagc aaagaaaata tcagaaacac acaactgaat gctgagattc
     2521 aagatttcaa tagcataaag aatagcccgc attgtgttga tctgctcgct tcgctagccc
     2581 ccgatttttt ccagaatggc gatgacccct catcggattt cgaatcagaa tatcgaaaaa
     2641 atatactgct aaaattgctt aatgtattgg atcttgcggg gaaaagaccc caacaccggg
     2701 ggcatcttgc tttcgtgaat tccatagcct atttaagaaa atggcttacc actacaatag
     2761 caacgcaaag tagagaatac ctctcatggt taaaaatgga acaaccttta tctgatgatc
     2821 ctctactaaa aaattttgct gatgtgttta tcccagtgag catacataat gagcctgcgc
     2881 tgatttgtct gttaaccgca gaatgtcaga agaaaaaggg ctttagtgat gcttatcagg
     2941 ccgcatttgc ctcccggcgt attgtcattg tgctgactca gactgcctct gcgacgaatg
     3001 gtgtgaatct ggatttcaac ctgcctgaat ctggcaaaaa tatggatctg acctgtttat
     3061 atttgctgga atcacggcat tattactttt ccgcttttca ggccaatgca gaaaacaatg
     3121 atgatatggc gcatgcgggt ttccagttac gcaacctgga aaaactgctc cgtgctggtg
     3181 aaatttctcg tcaacagcat caacgcttcc ttctgccatt aatgatgaac cgaaaatatg
     3241 agatttcccg attaaacggc gaatataaga gaaccagtga ttacataaaa aatacggcag
     3301 ctaatgtgca gcaacaggtc ggtcgtattg aacgagcctg gtgtgaagtc cccaatgctg
     3361 aaatccatct tgattctgaa ctggcaaaag acctttcgcg ctttgcttca ttgcctattt
     3421 atacatcgaa ccgtcgtcag ctttctgcgc tcaatcgcca gttactaaat attttacggg
     3481 aacgtgatga aaaaatgcag gacaatttcc ttaacctgat aatgacaccg acacaaccgg
     3541 gtaaactggt tttggattat atcgataaac gactggttcc ggccatacga ttgttgcgag
     3601 agcaatcaac acccagaaat gatgttacgc aactctggag acaattaggt aaagcgatac
     3661 ttcaacacga tcttgcctgg aaaccaggac ataatgactt tggtattgat cttgaactgt
     3721 acaagtgggc ttgttttaaa aaaccagaat ctgcatttaa ttgtggagca atctggtatt
     3781 cccctcagac ctggcagttt tttgaggaag cagcagaagg acggcgtaag tacaatcctc
     3841 agatgttgta tcaatatata cagcgtcatt cggacattac tgactggttt aaccgtaaag
     3901 ggtacagaac atctttaatg ccttacgcta acgatctgga agaacattac gccttccacc
     3961 cgctgatcgt acaacgtcta cttatggggc gcctgggtga agaggctatt cgggcactac
     4021 ttcatgagcg ttataaaatt attacaacga ctcaggtaag cgaccatcga attttcgaac
     4081 tgtatgattt tagtgtaaaa aactcagatt accgcataga tgcgaaattt tggggacagg
     4141 atactctcga caaagcggat gaagaatatc agcaatggct ggcgtctggc actgacccga
     4201 atcaaacccc tctgggatta agtagcaaat tagcaaaaat acgtgcgatt gagggagaca
     4261 acgtcaaact tgtaatcgct aatttcgtgg ctccccattc tgattgtcag ttgctgggat
     4321 tttcaagtca gttaattccg actcaggatc tttatcatgc agatatcctc atcctcggcg
     4381 ggtgtattac tcccgacaca gtttcatcag ttacactggg gtttgaaaat cttactacca
     4441 tgatctacca gaatattcat gaaagggcat gaatcggtac cctcgagaag cttgcggccg
     4501 cggggatcca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     4561 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     4621 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     4681 ggaggtgtgg gaggtttttt cggatcctct agagtcgatc tgcaggcatg ctagcttggc
     4741 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4801 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     4861 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4921 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4981 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5041 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5101 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5161 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5221 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5281 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5341 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5401 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     5461 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     5521 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     5581 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     5641 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     5701 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     5761 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     5821 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     5881 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     5941 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6001 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6061 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6121 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6181 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     6241 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     6301 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     6361 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     6421 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     6481 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     6541 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     6601 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     6661 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     6721 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     6781 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     6841 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     6901 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     6961 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     7021 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     7081 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     7141 actgagagtg caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     7201 catcaggcgc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     7261 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
     7321 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgagt tcgagcttgc
     7381 atgcctgcag gtcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     7441 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     7501 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     7561 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     7621 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     7681 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     7741 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     7801 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     7861 cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga accgtcagat
     7921 cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg accgatccag
     7981 cctccggact ctagaggatc cggtactaga ggaactgaaa aaccagaaag ttaactggta
     8041 agtttagtct ttttgtcttt tatttcaggt cccggatccg gtggtggtgc aaatcaaaga
     8101 actgctcctc agtggatgtt gcctttactt ctaggcctgt acggaagtgt tacttctgct
     8161 ctaaaagctg cggaattgta cccgcgggcc caccatggtg agcaagggcg aggagctgtt
     8221 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     8281 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     8341 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttga cctacggcgt
     8401 gcagt
//
