LOCUS Exported 7763 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION C-terminal EGFP tag. Gateway destination vector for mammalian expression.. ACCESSION . VERSION . KEYWORDS pDEST-CMV-C-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7763) AUTHORS Agrotis A, Pengo N, Burden JJ, Ketteler R TITLE Redundancy of human ATG4 protease isoforms in autophagy and LC3/GABARAP processing revealed in cells. JOURNAL Autophagy. 2019 Jan 21. doi: 10.1080/15548627.2019.1569925. PUBMED 30661429 REFERENCE 2 (bases 1 to 7763) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7763 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(46..65) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 237..616 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 617..820 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 771..791 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" protein_bind 890..1014 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 1039..1069 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 1123..1803 /codon_start=1 /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE DPAY" primer_bind complement(1189..1208) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 2123..2428 /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind 2307..2326 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(2469..2593) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" CDS 2610..3329 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 2610..3326 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2655..2676) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2916..2935) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3263..3284 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3356..3373) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3362..3586 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3632..4060 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3719..3738) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3929..3950 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4069..4089) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 4074..4403 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 4254..4389 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4316..4335 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4470..5264 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(4524..4543) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 5134..5153 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 5438..5559 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5475..5494) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5529..5548 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5608..5624) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5608..5624) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5621..5643) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5632..5648 /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(5656..5686) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5701..5722 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5839..5856) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(6010..6598) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(6090..6109) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6769..7629) /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 7392..7411 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7630..7734) /gene="bla" /label=AmpR promoter ORIGIN 1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga 61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg 121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct 181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca 241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata 301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga 361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt 421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt 481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca 541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt 601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt 661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca 721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg 781 cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga accgtcagaa 841 ttgatctggt accggcgtta acgacgctag cggagttaag cttgatcaaa caagtttgta 901 caaaaaagct gaacgagaaa cgtaaaatga tataaatatc aatatattaa attagatttt 961 gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac tatggcggcc 1021 gcattaggca ccccaggctt tacactttat gcttccggct cgtataatgt gtggattttg 1081 agttaggatc cgtcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact 1141 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag 1201 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag 1261 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg 1321 atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt gatatgggat 1381 agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg 1441 agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt 1501 tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca 1561 gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc 1621 ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg 1681 ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat 1741 gaattacaac agtactgcga tgagtggcag gcggggcgta atctagagga tccggcttac 1801 taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata agaatatata 1861 ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc gtattacagt 1921 gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa tatctccggt 1981 ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg 2041 gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct 2101 gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa gagagagccg 2161 ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt 2221 gatccccctg gccagtgcac gtctgctgtc agataaagtc ccccgtgaac tttacccggt 2281 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt 2341 ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc 2401 cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtctgc 2461 aggtcgacca tagtgactgg atatgttgtg ttttacagta ttatgtagtc tgttttttat 2521 gcaaaatcta atttaatata ttgatattta tatcatttta cgtttctcgt tcagctttct 2581 tgtacaaagt ggttcgatct agactcgaga tggtgagcaa gggcgaggag ctgttcaccg 2641 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 2701 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 2761 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt 2821 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 2881 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 2941 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 3001 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg 3061 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca 3121 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 3181 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag 3241 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 3301 ctctcggcat ggacgagctg tacaagtaat taattaagat aaacccgctg atcagcctcg 3361 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc 3421 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt 3481 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat 3541 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa 3601 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg 3661 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct 3721 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta 3781 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa 3841 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct 3901 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc 3961 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg 4021 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc 4081 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc 4141 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc 4201 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc 4261 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt 4321 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt 4381 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg 4441 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt 4501 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct 4561 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga 4621 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg 4681 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact 4741 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg 4801 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct 4861 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg 4921 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt 4981 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg 5041 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc 5101 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag 5161 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt 5221 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt 5281 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc 5341 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca 5401 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa 5461 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca 5521 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac 5581 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc 5641 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta 5701 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 5761 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 5821 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 5881 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 5941 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 6001 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 6061 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 6121 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 6181 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 6241 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 6301 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 6361 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 6421 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 6481 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 6541 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 6601 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 6661 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 6721 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 6781 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 6841 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat 6901 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 6961 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg 7021 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 7081 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 7141 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt 7201 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc 7261 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga 7321 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc 7381 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa 7441 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 7501 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 7561 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg 7621 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat 7681 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt 7741 tccccgaaaa gtgccacctg acg //