LOCUS       sequence-319251-        7658 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 7658)
  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 7658)
  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 GFP
FEATURES             Location/Qualifiers
     source          1..7658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        180..559
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        560..763
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     714..734
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     760..784
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          899..1031
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1014..1033
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             1100..1123
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             join(1133..1135,1136..1138,1139..1849)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1178..1199)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1439..1458)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1786..1807
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1868..1885
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     misc_feature    1929..2517
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1982..2002)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2400..2411)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2650..2784
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2687..2706)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2741..2760
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  2813..2978
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      3162..3616
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3249..3268)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3458..3479
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3642..3746
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3747..3815,3816..4607)
                     /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(3965..3984)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4778..5366
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5267..5286
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5520..5537
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  5671..5895
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(5962..6495)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(6684..6689,6690..6706,6707..6712))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    7229..7277
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        complement(7413..7522)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        7567..7658
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     7583..7600
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
ORIGIN
        1 attccggatt attcataccg tcccaccatc gggcgcggat cctcaatatt ggccattagc
       61 catattattc attggttata tagcataaat caatattggc tattggccat tgcatacgtt
      121 gtatctatat cataatatgt acatttatat tggctcatgt ccaatatgac cgccatgttg
      181 gcattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
      241 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      301 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      361 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      421 agtgtatcat atgccaagtc cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      481 gcattatgcc cagtacatga ccttacggga ctttcctact tggcagtaca tctacgtatt
      541 agtcatcgct attaccatgg tgatgcggtt ttggcagtac accaatgggc gtggatagcg
      601 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      661 gcaccaaaat caacgggact ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat
      721 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca
      781 gatcactaga agctttattg cggtagttta tcacagttaa attgctaacg cagtcagtgc
      841 ttctgacaca acagtctcga acttaagctg cagaagttgg tcgtgaggca ctgggcaggt
      901 aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct tgtcgagaca
      961 gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc actttgcctt
     1021 tctctccaca ggtgtccact cccagttcaa ttacagctct taagggatcc cggtccgaag
     1081 cgcgcggaat tccaccatgc accatcatca tcaccaccat cacgcggcag ccatggtgag
     1141 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     1201 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     1261 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     1321 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
     1381 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     1441 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     1501 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     1561 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     1621 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     1681 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     1741 cacccagtcc aagctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     1801 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt ccggactcag
     1861 atctggactg gtgccgcgag gtagcggcgg ccgctttcga atctagagcc tgcagtctcg
     1921 acaagcttaa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     1981 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     2041 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     2101 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     2161 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     2221 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     2281 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaagctgacg tcctttccat
     2341 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     2401 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     2461 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcctg
     2521 aagcttgtcg agaagtacta gaggatcata atcagccata ccacatttgt agaggtttta
     2581 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
     2641 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     2701 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     2761 aatgtatctt atcatgtctg gatctgatca ctgcttgagc ctaggagatc cgaaccagat
     2821 aagtgaaatc tagttccaaa ctattttgtc atttttaatt ttcgtattag cttacgacgc
     2881 tacacccagt tcccatctat tttgtcactc ttccctaaat aatccttaaa aactccattt
     2941 ccacccctcc cagttcccaa ctattttgtc cgcccacagc ggggcatttt tcttcctgtt
     3001 atgtttttaa tcaaacatcc tgccaactcc atgtgacaaa ccgtcatctt cggctacttt
     3061 ttctctgtca cagaatgaaa atttttctgt catctcttcg ttattaatgt ttgtaattga
     3121 ctgaatatca acgcttattt gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
     3181 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3241 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3301 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     3361 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     3421 ttttcgccct ttgacgttgg agtccacgtt cttaatagtg gactcttgtt ccaaactgga
     3481 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3541 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3601 ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3661 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3721 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3781 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3841 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3901 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3961 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     4021 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     4081 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     4141 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     4201 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     4261 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     4321 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     4381 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     4441 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4501 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4561 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     4621 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     4681 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     4741 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4801 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4861 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4921 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4981 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     5041 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     5101 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     5161 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     5221 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     5281 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     5341 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     5401 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     5461 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     5521 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     5581 tgtgcggtat ttcacaccgc atagaccagc cgcgtaacct ggcaaaatcg gttacggttg
     5641 agtaataaat ggatgccctg cgtaagcggg tgtgggcgga caataaagtc ttaaactgaa
     5701 caaaatagat ctaaactatg acaataaagt cttaaactag acagaatagt tgtaaactga
     5761 aatcagtcca gttatgctgt gaaaaagcat actggacttt tgttatggct aaagcaaact
     5821 cttcattttc tgaagtgcaa attgcccgtc gtattaaaga ggggcgtggc caagggcatg
     5881 gtaaagacta tattcgcggc gttgtgacaa tttaccgaac aactccgcgg ccgggaagcc
     5941 gatctcggct tgaacgaatt gttaggtggc ggtacttggg tcgatatcaa agtgcatcac
     6001 ttcttcccgt atgcccaact ttgtatagag agccactgcg ggatcgtcac cgtaatctgc
     6061 ttgcacgtag atcacataag caccaagcgc gttggcctca tgcttgagga gattgatgag
     6121 cgcggtggca atgccctgcc tccggtgctc gccggagact gcgagatcat agatatagat
     6181 ctcactacgc ggctgctcaa acttgggcag aacgtaagcc gcgagagcgc caacaaccgc
     6241 ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta cggagcaagt tcccgaggta
     6301 atcggagtcc ggctgatgtt gggagtaggt ggctacgtct ccgaactcac gaccgaaaag
     6361 atcaagagca gcccgcatgg atttgacttg gtcagggccg agcctacatg tgcgaatgat
     6421 gcccatactt gagccaccta actttgtttt agggcgactg ccctgctgcg taacatcgtt
     6481 gctgctgcgt aacatcgttg ctgctccata acatcaaaca tcgacccacg gcgtaacgcg
     6541 cttgctgctt ggatgcccga ggcatagact gtacaaaaaa acagtcataa caagccatga
     6601 aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa ggttctggac cagttgcgtg
     6661 atcgcatacg ctacttgcat tacagtttac gaaccgaaca ggcttatgtc aactgggttc
     6721 gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac cttgggcagc agcgaagtcg
     6781 aggcatttct gtcctggctg gcgaacgagc gcaaggtttc ggtctccacg catcgtcagg
     6841 cattggcggc cttgctgttc ttctacggca aggtgctgtg cacggatctg cccttgcttc
     6901 aggagatcgg tagacctcgg ccgtcgcggc gcttgccggt ggtgctgacc ccggatgaag
     6961 tggttcgcat cctcggtttt ctggaaggcg agcatcgttt gttcgcccag gactctagct
     7021 atagttctag tggttggcta cagctttgtt taaacaaagc tggctatggc agggcttgcc
     7081 gccccgacgt tggctgcgag ccctgggcct tcacccgaac ttgggggttg gggtggggaa
     7141 aaggaagaaa cgcgggcgta ttggtcccaa tggggtctcg gtggggtatc gacagagtgc
     7201 cagccctggg accgaacccc gcgtttatga acaaacgacc caacacccgt gcgttttatt
     7261 ctgtcttttt attgccgtca tagcgcgggt tccttccggt attgtctcat tccgtgtttc
     7321 agttagcctc ccccatctcc cggtaccgca tgctatgcat cagctgctag caccatggct
     7381 cgagatcccg ggtgatcaag tcttcgtcga gtgattgtaa ataaaatgta atttacagta
     7441 tagtatttta attaatatac aaatgatttg ataataattc ttatttaact ataatatatt
     7501 gtgttgggtt gaattaaagg tccgtatact agtatcgatt cgcgacctac tccggaatat
     7561 taatagatca tggagataat taaaatgata accatctcgc aaataaataa gtattttact
     7621 gttttcgtaa cagttttgta ataaaaaaac ctataaat
//
