LOCUS       sequence-318834-        7671 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 7671)
  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 7671)
  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 C term GFP His8
FEATURES             Location/Qualifiers
     source          1..7671
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          720..852
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     835..854
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             964..981
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             join(994..996,997..1707)
                     /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(1036..1057)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1297..1316)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1644..1665
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1723..1746
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     misc_feature    1763..2351
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1816..1836)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2234..2245)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2484..2618
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2521..2540)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2575..2594
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  2647..2812
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      2996..3450
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3083..3102)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3292..3313
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3476..3580
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3581..3649,3650..4441)
                     /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(3799..3818)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4612..5200
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5101..5120
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5354..5371
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  5505..5729
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(5796..6329)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(6518..6523,6524..6540,6541..6546))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    7063..7111
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     7136..7155
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     promoter        complement(7247..7356)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        7401..7492
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     7417..7434
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcactag aagctttatt gcggtagttt atcacagtta aattgctaac gcagtcagtg
      661 cttctgacac aacagtctcg aacttaagct gcagaagttg gtcgtgaggc actgggcagg
      721 taagtatcaa ggttacaaga caggtttaag gagaccaata gaaactgggc ttgtcgagac
      781 agagaagact cttgcgtttc tgataggcac ctattggtct tactgacatc cactttgcct
      841 ttctctccac aggtgtccac tcccagttca attacagctc ttaagggatc ccggtccgaa
      901 gcgcgcggaa ttcaaaggcc tacgtcgacg agctcactag tcgcggccgc tttcgaatct
      961 agactggtgc cgcgcggctc tgccgccgcc gcggtgagca agggcgagga gctgttcacc
     1021 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     1081 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
     1141 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     1201 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     1261 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     1321 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     1381 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     1441 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     1501 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     1561 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa gctgagcaaa
     1621 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     1681 actctcggca tggacgagct gtacaagtcc ggactcagat ctcaccatca tcatcaccac
     1741 catcactgat ctcgacaagc ttaatcaacc tctggattac aaaatttgtg aaagattgac
     1801 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     1861 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     1921 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     1981 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     2041 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     2101 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaagct
     2161 gacgtccttt ccatggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     2221 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     2281 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     2341 cgcctccccg cctgaagctt gtcgagaagt actagaggat cataatcagc cataccacat
     2401 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
     2461 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
     2521 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     2581 tgtccaaact catcaatgta tcttatcatg tctggatctg atcactgctt gagcctagga
     2641 gatccgaacc agataagtga aatctagttc caaactattt tgtcattttt aattttcgta
     2701 ttagcttacg acgctacacc cagttcccat ctattttgtc actcttccct aaataatcct
     2761 taaaaactcc atttccaccc ctcccagttc ccaactattt tgtccgccca cagcggggca
     2821 tttttcttcc tgttatgttt ttaatcaaac atcctgccaa ctccatgtga caaaccgtca
     2881 tcttcggcta ctttttctct gtcacagaat gaaaattttt ctgtcatctc ttcgttatta
     2941 atgtttgtaa ttgactgaat atcaacgctt atttgcagcc tgaatggcga atgggacgcg
     3001 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     3061 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     3121 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     3181 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     3241 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttcttaat agtggactct
     3301 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     3361 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     3421 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     3481 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     3541 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     3601 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     3661 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     3721 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     3781 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     3841 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     3901 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     3961 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     4021 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     4081 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     4141 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     4201 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4261 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     4321 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     4381 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     4441 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     4501 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     4561 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4621 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     4681 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     4741 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     4801 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     4861 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     4921 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     4981 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5041 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5101 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5161 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5221 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5281 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     5341 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     5401 tctccttacg catctgtgcg gtatttcaca ccgcatagac cagccgcgta acctggcaaa
     5461 atcggttacg gttgagtaat aaatggatgc cctgcgtaag cgggtgtggg cggacaataa
     5521 agtcttaaac tgaacaaaat agatctaaac tatgacaata aagtcttaaa ctagacagaa
     5581 tagttgtaaa ctgaaatcag tccagttatg ctgtgaaaaa gcatactgga cttttgttat
     5641 ggctaaagca aactcttcat tttctgaagt gcaaattgcc cgtcgtatta aagaggggcg
     5701 tggccaaggg catggtaaag actatattcg cggcgttgtg acaatttacc gaacaactcc
     5761 gcggccggga agccgatctc ggcttgaacg aattgttagg tggcggtact tgggtcgata
     5821 tcaaagtgca tcacttcttc ccgtatgccc aactttgtat agagagccac tgcgggatcg
     5881 tcaccgtaat ctgcttgcac gtagatcaca taagcaccaa gcgcgttggc ctcatgcttg
     5941 aggagattga tgagcgcggt ggcaatgccc tgcctccggt gctcgccgga gactgcgaga
     6001 tcatagatat agatctcact acgcggctgc tcaaacttgg gcagaacgta agccgcgaga
     6061 gcgccaacaa ccgcttcttg gtcgaaggca gcaagcgcga tgaatgtctt actacggagc
     6121 aagttcccga ggtaatcgga gtccggctga tgttgggagt aggtggctac gtctccgaac
     6181 tcacgaccga aaagatcaag agcagcccgc atggatttga cttggtcagg gccgagccta
     6241 catgtgcgaa tgatgcccat acttgagcca cctaactttg ttttagggcg actgccctgc
     6301 tgcgtaacat cgttgctgct gcgtaacatc gttgctgctc cataacatca aacatcgacc
     6361 cacggcgtaa cgcgcttgct gcttggatgc ccgaggcata gactgtacaa aaaaacagtc
     6421 ataacaagcc atgaaaaccg ccactgcgcc gttaccaccg ctgcgttcgg tcaaggttct
     6481 ggaccagttg cgtgatcgca tacgctactt gcattacagt ttacgaaccg aacaggctta
     6541 tgtcaactgg gttcgtgcct tcatccgttt ccacggtgtg cgtcacccgg caaccttggg
     6601 cagcagcgaa gtcgaggcat ttctgtcctg gctggcgaac gagcgcaagg tttcggtctc
     6661 cacgcatcgt caggcattgg cggccttgct gttcttctac ggcaaggtgc tgtgcacgga
     6721 tctgcccttg cttcaggaga tcggtagacc tcggccgtcg cggcgcttgc cggtggtgct
     6781 gaccccggat gaagtggttc gcatcctcgg ttttctggaa ggcgagcatc gtttgttcgc
     6841 ccaggactct agctatagtt ctagtggttg gctacagctt tgtttaaaca aagctggcta
     6901 tggcagggct tgccgccccg acgttggctg cgagccctgg gccttcaccc gaacttgggg
     6961 gttggggtgg ggaaaaggaa gaaacgcggg cgtattggtc ccaatggggt ctcggtgggg
     7021 tatcgacaga gtgccagccc tgggaccgaa ccccgcgttt atgaacaaac gacccaacac
     7081 ccgtgcgttt tattctgtct ttttattgcc gtcatagcgc gggttccttc cggtattgtc
     7141 tccttccgtg tttcagttag cctcccccat ctcccggtac cgcatgctat gcatcagctg
     7201 ctagcaccat ggctcgagat cccgggtgat caagtcttcg tcgagtgatt gtaaataaaa
     7261 tgtaatttac agtatagtat tttaattaat atacaaatga tttgataata attcttattt
     7321 aactataata tattgtgttg ggttgaatta aaggtccgta tactagtatc gattcgcgac
     7381 ctactccgga atattaatag atcatggaga taattaaaat gataaccatc tcgcaaataa
     7441 ataagtattt tactgttttc gtaacagttt tgtaataaaa aaacctataa atattccgga
     7501 ttattcatac cgtcccacca tcgggcgcgg atcctcaata ttggctatta gccatattat
     7561 tcattggtta tatagcataa atcaatattg gctattggcc attgcatacg ttgtatctat
     7621 atcataatat gtacatttat attggctcat gtccaatatg accgccatgt t
//
