LOCUS       sequence-209763-        4643 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of E. coli LF82_781 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4643)
  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 4643)
  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_781
FEATURES             Location/Qualifiers
     source          1..4643
                     /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    1799..1933
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1823..1842)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1877..1896
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2044..2060)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2044..2060)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2057..2079)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2068..2084
                     /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(2092..2098,2099..2116,2117..2122))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2137..2158
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2275..2292)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2446..3034)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2526..2545)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3205..3996,3997..4065))
                     /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     3828..3847
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4066..4170)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4238..4256
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4294..4316)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4416..4435
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(4629..4643,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(4643,1..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 aacttattac agccattggc ggcacgcgtt gctaattcac gatggtgatt
     1681 ttatttccac gcaaacggtt tcgtcagcgc atcagattct ttataatgac gcccgtttcc
     1741 cccccttggg tacaccgaaa gcttgaatcg gtaccctcga gaagcttgcg gccgcgggga
     1801 tccagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     1861 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     1921 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt
     1981 gtgggaggtt ttttcggatc ctctagagtc gatctgcagg catgctagct tggcgtaatc
     2041 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     2101 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     2161 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     2221 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     2281 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     2341 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     2401 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     2461 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     2521 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     2581 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     2641 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     2701 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     2761 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     2821 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     2881 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     2941 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     3001 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     3061 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     3121 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     3181 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     3241 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     3301 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     3361 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     3421 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     3481 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     3541 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     3601 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     3661 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     3721 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     3781 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     3841 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     3901 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     3961 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     4021 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     4081 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     4141 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     4201 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     4261 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     4321 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     4381 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
     4441 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     4501 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     4561 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     4621 agggttttcc cagtcacgac gtt
//
