LOCUS Exported 7776 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synchronize trafficking of TNF from the ER (RUSH system). ACCESSION . VERSION . KEYWORDS Ii-Str_TNF-SBP-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7776) AUTHORS Boncompain G, Divoux S, Gareil N, de Forges H, Lescure A, Latreche L, Mercanti V, Jollivet F, Raposo G, Perez F TITLE Synchronization of secretory protein traffic in populations of cells. JOURNAL Nat Methods. 2012 Mar 11;9(5):493-8. doi: 10.1038/nmeth.1928. PUBMED 22406856 REFERENCE 2 (bases 1 to 7776) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7776 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 61..364 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 365..568 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 519..539 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" CDS 1401..1427 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" intron 1946..2175 /label=chimeric intron /note="chimera between introns from adenovirus and immunoglobulin heavy chain genes" primer_bind 2158..2177 /label=pMT2-F /note="Synthetic intron, forward primer" misc_feature 2233..2806 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(2400..2417) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 2627..2646 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 3577..3687 /codon_start=1 /product="streptavidin-binding peptide" /label=SBP /note="selected from a peptide library; binds streptavidin with nanomolar affinity (Keefe et al., 2001)" /translation="DEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP" CDS 3697..4410 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" CDS 3697..4410 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(3739..3760) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(4000..4019) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 4347..4368 /label=EGFP-C /note="EGFP, forward primer" polyA_signal 4564..4685 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4601..4620) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4655..4674 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(4692..5147) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4829..4850) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5041..5060 /label=F1ori-R /note="F1 origin, reverse primer" promoter 5174..5278 /gene="bla" /label=AmpR promoter promoter 5280..5637 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(5303..5323) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 5488..5623 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5550..5569 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 5672..6466 /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 GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(5726..5745) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 6336..6355 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(6654..6673) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 6698..6745 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 7074..7662 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7563..7582 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg 61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta 601 ccggtcgtac gtctccggat tcgaattaat tcggatccgt gatgcacagg aggagagcca 661 gggcctgtcg ggaagatcaa aagccagtca tcgatgatca gcgcgacctt atctccaaca 721 atgagcaact gcccatgctg ggccggcgcc ctggggcccc ggagagcaag tgcagccgcg 781 gagccctgta cacaggcttt tccatcctgg tgactctgct cctcgctggc caggccacca 841 ccgcctactt cctgtaccag cagcagggcc ggctggacaa actgacagtc acctcccaga 901 acctgcagct cgagatggac cctagcaaag actcaaaagc tcaggtgtcc gctgccgagg 961 ctggcattac tggaacatgg tacaatcagc tcgggagcac ctttattgtg actgctggag 1021 ccgatggagc cctcaccgga acatacgaat ctgctgtggg aaacgccgaa tcacggtacg 1081 tcctcactgg ccgatacgat agtgcccctg ccaccgacgg atctgggact gccctgggat 1141 ggactgtcgc ttggaaaaac aactaccgga atgctcattc tgccacaaca tggagtggac 1201 agtacgtggg aggcgctgag gctagaatca atacacagtg gctgctcaca tctggcacaa 1261 ccgaggcaaa tgcttggaaa tccaccctgg tgggacatga cacattcacc aaagtgaaac 1321 cctccgccgc ttcaatcgat gccgccaaaa aagccggagt caacaacggc aatcctctgg 1381 atgccgtcca gcaggtcgac tatccgtacg acgtaccaga ctacgcactc gacctgcagc 1441 tggagaacct gcgcatgaag cttcccaagc ctcccaagcc tgtgagcaag atgcgcatgg 1501 ccaccccgct gctgatgcat gcgctgccca tgggagccct gccccagggg cccatgcaga 1561 atgccaccaa gtatggcaac atgacagagg accatgtgat gcacctgctc cagaatgctg 1621 accccctgaa ggtgtacccg ccactgaagg ggagcttccc ggagaacctg agacacctta 1681 agaacaccat ggagaccata gactggaagg tctttgagag ctggatgcac cattggctcc 1741 tgtttgaaat gagcaggcac tccttggagc aaaagcccac tgacgctcca ccgaaagagt 1801 cactggaact ggaggacccg tcttctgggc tgggtgtgac caagcaggat ctgggcccag 1861 tccccatgtg aggatccgcg gccgcataga taactgatcc agtgtgctgg aattaattcg 1921 ctgtctgcga gggccagctg ttggggtgag tactccctct caaaagcggg catgacttct 1981 gcgctaagat tgtcagtttc caaaaacgag gaggatttga tattcacctg gcccgcggtg 2041 atgcctttga gggtggccgc gtccatctgg tcagaaaaga caatcttttt gttgtcaagc 2101 ttgaggtgtg gcaggcttga gatctggcca tacacttgag tgacaatgac atccactttg 2161 cctttctctc cacaggtgtc cactcccagg tccaactgca ggtcgagcat gcatctaggg 2221 cggccaattc cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga 2281 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa 2341 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc 2401 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc 2461 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg 2521 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca 2581 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag 2641 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct 2701 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc 2761 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataagctt gccacaaccc 2821 gggaggcgcg ccatgtctac cgagtctatg attagggacg tggaactggc tgaggaggca 2881 ctgcccaaaa aaaccggcgg accacagggc tctaggagat gtctgtttct gtctctgttc 2941 tcttttctca tcgtggctgg cgctacaaca ctcttctgtc tgctccattt cggcgtgatt 3001 ggaccacagc gagaggaatt tccccgggat ctgtcactca tttcaccact ggcacaggct 3061 gtccgatctt catctcggac tccatccgac aaacctgtcg cccatgtcgt cgccaaccca 3121 caggccgagg gccagctcca gtggctcaat aggagggcaa acgctctgct cgccaatggc 3181 gtggaactcc gggataacca gctcgtcgtg cctagtgagg gactgtacct catctactcc 3241 caggtgctgt ttaagggcca gggatgtcct tctacacatg tgctgctcac acacacaatt 3301 tcacggatcg ccgtgtctta ccagactaaa gtcaatctgc tctctgccat caaatcccca 3361 tgtcagcggg aaacacctga gggcgctgag gctaaacctt ggtacgaacc catctacctc 3421 ggaggcgtgt tccagctgga gaagggcgat agactgagtg ctgagatcaa tcgacccgac 3481 tacctcgact ttgccgaatc tggccaggtc tactttggca tcattgctct gggcctcgag 3541 ggccgagctc atggcgcacc taggcctttg aattccgacg agaagaccac tggttggcga 3601 ggtggacacg ttgttgaagg actggctggg gaacttgaac aacttcgtgc acgactggag 3661 catcacccac aaggtcaacg tgaaccacct gcaggtgtga gcaagggcga ggagctgttc 3721 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc 3781 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc 3841 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg 3901 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg 3961 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc 4021 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc 4081 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac 4141 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc 4201 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc 4261 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc 4321 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg 4381 atcactctcg gcatggacga gctgtacaag ggccggcctt aaggcctcga gggccaatta 4441 attaactcta gataactgat cataatcagc cataccacat ttgtagaggt tttacttgct 4501 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt 4561 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc 4621 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta 4681 tcttaacgcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta 4741 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga 4801 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa 4861 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga 4921 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc 4981 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga 5041 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg 5101 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt 5161 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta 5221 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc 5281 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 5341 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 5401 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 5461 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 5521 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc 5581 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat 5641 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg 5701 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg 5761 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa 5821 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct 5881 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga 5941 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc 6001 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac 6061 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc 6121 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact 6181 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga 6241 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg 6301 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga 6361 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga 6421 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg 6481 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc 6541 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc 6601 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca 6661 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa 6721 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct 6781 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca 6841 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc 6901 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa 6961 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt 7021 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat 7081 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg 7141 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga 7201 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac 7261 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt 7321 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag 7381 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc 7441 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag 7501 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca 7561 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt 7621 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc 7681 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc 7741 cctgattctg tggataaccg tattaccgcc atgcat //