LOCUS       sequence-319254-        7695 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector optimized for use in screening assays, as well as for 
            efficient production of baculovirus and robust expression of the 
            target protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7695)
  AUTHORS   Goehring A, Lee CH, Wang KH, Michel JC, Claxton DP, Baconguis I, 
            Althoff T, Fischer S, Garcia KC, Gouaux E
  TITLE     Screening and large-scale expression of membrane proteins in 
            mammalian cells for structural studies.
  JOURNAL   Nat Protoc. 2014 Nov;9(11):2574-85. doi: 10.1038/nprot.2014.173. 
            Epub 2014 Oct 9.
  PUBMED    25299155
REFERENCE   2  (bases 1 to 7695)
  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"; doi: 
            "10.1038/nprot.2014.173"; journalName: "Nat Protoc"; date: 
            "2014-11"; volume: "9"; issue: "11"; pages: "2574-85"
COMMENT     Sequence Label: pEG BacMam N term His8 eGFP 3C
FEATURES             Location/Qualifiers
     source          1..7695
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     201..225
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          340..472
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     455..474
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             541..564
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             join(574..576,577..1287)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(616..637)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(877..896)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1224..1245
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1306..1329
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     misc_feature    1407..1995
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1460..1480)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1878..1889)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2128..2262
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2165..2184)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2219..2238
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  2291..2456
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      2640..3094
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2727..2746)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2936..2957
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3120..3224
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3225..3293,3294..4085)
                     /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     complement(3443..3462)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4256..4844
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4745..4764
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4998..5015
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  5149..5373
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(5440..5973)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(6162..6167,6168..6184,6185..6190))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    6707..6755
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     6780..6799
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     promoter        complement(6891..7000)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        7045..7136
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     7061..7078
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     enhancer        7316..7695
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta caccaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctcgttta gtgaaccgtc agatcactag aagctttatt
      241 gcggtagttt atcacagtta aattgctaac gcagtcagtg cttctgacac aacagtctcg
      301 aacttaagct gcagaagttg gtcgtgaggc actgggcagg taagtatcaa ggttacaaga
      361 caggtttaag gagaccaata gaaactgggc ttgtcgagac agagaagact cttgcgtttc
      421 tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac aggtgtccac
      481 tcccagttca attacagctc ttaagggatc ccggtccgaa gcgcgcggaa ttacaccatg
      541 caccatcatc atcaccacca tcacggttca ggagtgagca agggcgagga gctgttcacc
      601 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
      661 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
      721 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
      781 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
      841 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
      901 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
      961 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     1021 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     1081 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     1141 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa gctgagcaaa
     1201 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     1261 actctcggca tggacgagct gtacaagtcc ggactcagat ctggactgga agttctgttc
     1321 cagggtcctg aattcaaagg cctacgtcga cgagctcact agtcgcggcc gctttcgaat
     1381 ctagagcctg cagtctcgac aagcttaatc aacctctgga ttacaaaatt tgtgaaagat
     1441 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     1501 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     1561 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     1621 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     1681 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     1741 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     1801 agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
     1861 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     1921 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     1981 gggccgcctc cccgcctgaa gcttgtcgag aagtactaga ggatcataat cagccatacc
     2041 acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa
     2101 cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa
     2161 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     2221 ggtttgtcca aactcatcaa tgtatcttat catgtctgga tctgatcact gcttgagcct
     2281 aggagatccg aaccagataa gtgaaatcta gttccaaact attttgtcat ttttaatttt
     2341 cgtattagct tacgacgcta cacccagttc ccatctattt tgtcactctt ccctaaataa
     2401 tccttaaaaa ctccatttcc acccctccca gttcccaact attttgtccg cccacagcgg
     2461 ggcatttttc ttcctgttat gtttttaatc aaacatcctg ccaactccat gtgacaaacc
     2521 gtcatcttcg gctacttttt ctctgtcaca gaatgaaaat ttttctgtca tctcttcgtt
     2581 attaatgttt gtaattgact gaatatcaac gcttatttgc agcctgaatg gcgaatggga
     2641 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2701 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2761 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2821 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2881 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct taatagtgga
     2941 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3001 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3061 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc
     3121 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     3181 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     3241 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     3301 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     3361 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     3421 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     3481 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     3541 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     3601 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     3661 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     3721 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     3781 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     3841 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     3901 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     3961 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     4021 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     4081 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     4141 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     4201 atgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     4261 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     4321 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     4381 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     4441 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     4501 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     4561 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     4621 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     4681 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     4741 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     4801 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     4861 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     4921 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     4981 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt
     5041 attttctcct tacgcatctg tgcggtattt cacaccgcat agaccagccg cgtaacctgg
     5101 caaaatcggt tacggttgag taataaatgg atgccctgcg taagcgggtg tgggcggaca
     5161 ataaagtctt aaactgaaca aaatagatct aaactatgac aataaagtct taaactagac
     5221 agaatagttg taaactgaaa tcagtccagt tatgctgtga aaaagcatac tggacttttg
     5281 ttatggctaa agcaaactct tcattttctg aagtgcaaat tgcccgtcgt attaaagagg
     5341 ggcgtggcca agggcatggt aaagactata ttcgcggcgt tgtgacaatt taccgaacaa
     5401 ctccgcggcc gggaagccga tctcggcttg aacgaattgt taggtggcgg tacttgggtc
     5461 gatatcaaag tgcatcactt cttcccgtat gcccaacttt gtatagagag ccactgcggg
     5521 atcgtcaccg taatctgctt gcacgtagat cacataagca ccaagcgcgt tggcctcatg
     5581 cttgaggaga ttgatgagcg cggtggcaat gccctgcctc cggtgctcgc cggagactgc
     5641 gagatcatag atatagatct cactacgcgg ctgctcaaac ttgggcagaa cgtaagccgc
     5701 gagagcgcca acaaccgctt cttggtcgaa ggcagcaagc gcgatgaatg tcttactacg
     5761 gagcaagttc ccgaggtaat cggagtccgg ctgatgttgg gagtaggtgg ctacgtctcc
     5821 gaactcacga ccgaaaagat caagagcagc ccgcatggat ttgacttggt cagggccgag
     5881 cctacatgtg cgaatgatgc ccatacttga gccacctaac tttgttttag ggcgactgcc
     5941 ctgctgcgta acatcgttgc tgctgcgtaa catcgttgct gctccataac atcaaacatc
     6001 gacccacggc gtaacgcgct tgctgcttgg atgcccgagg catagactgt acaaaaaaac
     6061 agtcataaca agccatgaaa accgccactg cgccgttacc accgctgcgt tcggtcaagg
     6121 ttctggacca gttgcgtgat cgcatacgct acttgcatta cagtttacga accgaacagg
     6181 cttatgtcaa ctgggttcgt gccttcatcc gtttccacgg tgtgcgtcac ccggcaacct
     6241 tgggcagcag cgaagtcgag gcatttctgt cctggctggc gaacgagcgc aaggtttcgg
     6301 tctccacgca tcgtcaggca ttggcggcct tgctgttctt ctacggcaag gtgctgtgca
     6361 cggatctgcc cttgcttcag gagatcggta gacctcggcc gtcgcggcgc ttgccggtgg
     6421 tgctgacccc ggatgaagtg gttcgcatcc tcggttttct ggaaggcgag catcgtttgt
     6481 tcgcccagga ctctagctat agttctagtg gttggctaca gctttgttta aacaaagctg
     6541 gctatggcag ggcttgccgc cccgacgttg gctgcgagcc ctgggccttc acccgaactt
     6601 gggggttggg gtggggaaaa ggaagaaacg cgggcgtatt ggtcccaatg gggtctcggt
     6661 ggggtatcga cagagtgcca gccctgggac cgaaccccgc gtttatgaac aaacgaccca
     6721 acacccgtgc gttttattct gtctttttat tgccgtcata gcgcgggttc cttccggtat
     6781 tgtctccttc cgtgtttcag ttagcctccc ccatctcccg gtaccgcatg ctatgcatca
     6841 gctgctagca ccatggctcg agatcccggg tgatcaagtc ttcgtcgagt gattgtaaat
     6901 aaaatgtaat ttacagtata gtattttaat taatatacaa atgatttgat aataattctt
     6961 atttaactat aatatattgt gttgggttga attaaaggtc cgtatactag tatcgattcg
     7021 cgacctactc cggaatatta atagatcatg gagataatta aaatgataac catctcgcaa
     7081 ataaataagt attttactgt tttcgtaaca gttttgtaat aaaaaaacct ataaatattc
     7141 cggattattc ataccgtccc accatcgggc gcggatcctc aatattggcc attagccata
     7201 ttattcattg gttatatagc ataaatcaat attggctatt ggccattgca tacgttgtat
     7261 ctatatcata atatgtacat ttatattggc tcatgtccaa tatgaccgcc atgttggcat
     7321 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     7381 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     7441 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     7501 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     7561 tatcatatgc caagtccgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     7621 tatgcccagt acatgacctt acgggacttt cctacttggc agtacatcta cgtattagtc
     7681 atcgctatta ccatg
//
