LOCUS pEG BacMam N term StrepII eGFP 3C 7695 bp ds-DNA circular SYN 12-MAY-2021
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 pEG BacMam N term StrepII eGFP 3C
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 May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7695
/organism="synthetic DNA construct"
/mol_type="other DNA"
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="peptide that binds Strep-Tactin(R), an engineered
form of streptavidin"
/label=Strep-Tag II
/translation="WSHPQFEK"
CDS 574..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"
/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 3225..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"
/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(6162..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 tggagccacc cgcagttcga aaaaggttca 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
//
