LOCUS       sequence-209949-        4694 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of E. coli LF82_146 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4694)
  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 4694)
  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_146
FEATURES             Location/Qualifiers
     source          1..4694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     88..110
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     102..119
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     103..119
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        146..449
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        450..653
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     604..624
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     650..674
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          791..887
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             join(948..950,951..953,954..1664)
                     /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(993..1014)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1254..1273)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1601..1622
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1952..2086
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1976..1995)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2030..2049
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2197..2213)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2197..2213)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2210..2232)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2221..2237
                     /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(2245..2251,2252..2269,2270..2275))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2290..2311
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2428..2445)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2599..3187)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2679..2698)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3358..4149,4150..4218))
                     /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     3981..4000
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4219..4323)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4391..4409
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4447..4469)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4569..4588
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc
       61 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
      121 agttcgagct tgcatgcctg caggtcgtta cataacttac ggtaaatggc ccgcctggct
      181 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      241 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      301 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      361 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      421 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      481 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      541 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      601 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
      661 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc
      721 gggaccgatc cagcctccgg actctagagg atccggtact agaggaactg aaaaaccaga
      781 aagttaactg gtaagtttag tctttttgtc ttttatttca ggtcccggat ccggtggtgg
      841 tgcaaatcaa agaactgctc ctcagtggat gttgccttta cttctaggcc tgtacggaag
      901 tgttacttct gctctaaaag ctgcggaatt gtacccgcgg gcccaccatg gtgagcaagg
      961 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     1021 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     1081 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccacct
     1141 tgacctacgg cgtgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct
     1201 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
     1261 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     1321 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     1381 actacaacag ccacaaggtc tatatcaccg ccgacaagca gaagaacggc atcaaggtga
     1441 acttcaagac ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
     1501 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc
     1561 agtccaagct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     1621 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagcgaatt ggtggagggg
     1681 gcggttccgg tggcggaggt tcccgaattc gtcgacggag atctgatatg cacaaagcat
     1741 ctcccgttga gttaagaacg agtatcgaga tggcacatag cctcgctcaa attggagtca
     1801 ggtttgtgcc aataccagta gaaacagacg aagaatttca tacgttagcc acatcccttt
     1861 cacaaaagct ggaaatgatg gtggcgaaag cagaagcaga tgagagagac caggtatgaa
     1921 tcggtaccct cgagaagctt gcggccgcgg ggatccagac atgataagat acattgatga
     1981 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     2041 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     2101 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttcgg atcctctaga
     2161 gtcgatctgc aggcatgcta gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     2221 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     2281 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     2341 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     2401 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     2461 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     2521 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2581 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2641 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     2701 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     2761 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     2821 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     2881 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     2941 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3001 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     3061 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3121 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3181 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3241 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     3301 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     3361 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3421 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     3481 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     3541 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     3601 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     3661 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     3721 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     3781 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     3841 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     3901 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     3961 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     4021 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     4081 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     4141 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     4201 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     4261 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     4321 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     4381 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     4441 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     4501 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     4561 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     4621 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     4681 ctgttgggaa gggc
//
