LOCUS       sequence-209955-        5627 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of E. coli LF82_467 CDS fused to mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5627)
  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 5627)
  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_467
FEATURES             Location/Qualifiers
     source          1..5627
                     /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    2783..2917
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2807..2826)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2861..2880
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3028..3044)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3028..3044)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3041..3063)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3052..3068
                     /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(3076..3082,3083..3100,3101..3106))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3121..3142
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3259..3276)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3430..4018)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3510..3529)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4189..4980,4981..5049))
                     /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     4812..4831
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5050..5154)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5222..5240
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5278..5300)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5400..5419
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(5613..5627,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(5627,1..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 ctgatatgtt gacaaagctt accgtcgaaa cggcgctcaa tgcagaactg actgaccatc
     1681 tcggtcatga gaaaaatgct cctaaatcag gctctaatac ccgtaatggc tactcgtcaa
     1741 aaacggtatt gtgcgatgat ggcgagatcg aacttaacac gccgcgtgac cgtgaaaata
     1801 ccttcgaacc gcaactaatt aagaagcatc agacacgcat cacccagatg gacagccaga
     1861 ttttgtccct gtatgccaaa ggcatgacta cgcgggaaat cgttgccacg ttcaaagaaa
     1921 tgtacgacgc cgatgtctcg cccacgctga tatccaaagt gaccgatgcg gttaaagaac
     1981 aggtctctga atggcaaaat cgaccgttgg atgcactgta tcccattgtt tatcttgact
     2041 gtattgtggt taaggttcgt cacggcggga gtgtcattaa caaagcggtg ttcctcgcgc
     2101 tgggcatcaa taccgacggt cagaaagagc tgctgggtat gtggctggca gaaaatgaag
     2161 gtgcaaagtt ctggctgggt gtactgactg agctgaaaaa tcgtggcctt caggacatcc
     2221 tgattgcctg cgtggacggc ctgaaaggct tcccggaagc gataaatagc gtgtacccgc
     2281 agacgcatat tcagctctgc atcatccata tggtgcgcaa cagcctgaaa ttcgtgtcgt
     2341 ggaaggatta caaagccgtc accagcgggc tgaaagcggt atatcaggcc ccgacagagg
     2401 aagcggcgtt gatggcgttg gataagtttg caggcgtctg ggatgaaaaa tacccgcaaa
     2461 tcagcaagag ctggcgcaca cactgggaaa atctcaacac cttcttcggt tacccgccgg
     2521 atatccgcaa ggccatctac acgacgaatg ccatcgaatc gctgaacagc gtcattcggc
     2581 aggcgataaa gaagcgcaag gtgttcccaa cggacgactc ggtgcgaaaa gtgatttatc
     2641 tggcgataca gtcggcgtca aaaaaatgga gtatgccaat ccagaactgg cggctggcaa
     2701 tgagccgttt tattatcgag tttggtgacc gccttagcga gcacctttga atcggtaccc
     2761 tcgagaagct tgcggccgcg gggatccaga catgataaga tacattgatg agtttggaca
     2821 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     2881 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     2941 tatgtttcag gttcaggggg aggtgtggga ggttttttcg gatcctctag agtcgatctg
     3001 caggcatgct agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3061 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     3121 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3181 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3241 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3301 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3361 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3421 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3481 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3541 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3601 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3661 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3721 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3781 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     3841 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     3901 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     3961 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4021 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4081 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4141 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4201 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4261 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4321 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     4381 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4441 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4501 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4561 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4621 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4681 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     4741 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     4801 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     4861 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     4921 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     4981 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5041 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5101 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     5161 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     5221 aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct
     5281 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     5341 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc
     5401 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     5461 cgtaaggaga aaataccgca tcaggcgcca ttcgccattc aggctgcgca actgttggga
     5521 agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc
     5581 aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgtt
//
