LOCUS Exported 5655 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses GFP tagged Tac mutant in mammalian cells. The five O-glycosylation sites in the stem region are mutated, and the stem region of CD8a with mutations is inserted between GFP and Tac(5A).. ACCESSION . VERSION . KEYWORDS GFP-Cstem(7A)-Tac(5A) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5655) AUTHORS Sun X, Tie HC, Chen B, Lu L TITLE Glycans function as a Golgi export signal to promote the constitutive exocytic trafficking. JOURNAL J Biol Chem. 2020 Aug 21. pii: RA120.014476. doi: 10.1074/jbc.RA120.014476. PUBMED 32826314 REFERENCE 2 (bases 1 to 5655) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5655 /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" regulatory 597..606 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 672..1388 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 672..1388 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(717..738) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(978..997) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1325..1346 /label=EGFP-C /note="EGFP, forward primer" polyA_signal 2443..2564 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2480..2499) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2534..2553 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2571..3026) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2708..2729) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2920..2939 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3053..3157 /gene="bla" /label=AmpR promoter promoter 3159..3516 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3182..3202) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3367..3502 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3429..3448 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3551..4345 /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(3605..3624) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4215..4234 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4533..4552) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4577..4624 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4953..5541 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5442..5461 /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 gctagcgcca 601 ccatggattc atacctgctg atgtggggac tgctcacgtt catcatggtg cctggctgcc 661 aggcaaccgg tatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg 721 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg 781 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc 841 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg 901 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc 961 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg 1021 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca 1081 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca 1141 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg 1201 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc 1261 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg 1321 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc 1381 tgtacaagct cgagccagcg aagcccgcag cagctccagc gccgcgacca ccagcaccgg 1441 cgcccgcaat cgcggcacag cccctggcac tgcgcccaga ggcgtgtcgg ccagcggcgg 1501 ggggcgcagt gcacacgagg gggctggagc tctgtgacga tgacccgcca gagatcccac 1561 acgccacatt caaagccatg gcctacaagg aaggaaccat gttgaactgt gaatgcaaga 1621 gaggtttccg cagaataaaa agcgggtcac tctatatgct ctgtacagga aactctagcc 1681 actcgtcctg ggacaaccaa tgtcaatgca caagctctgc cactcggaac acaacgaaac 1741 aagtgacacc tcaacctgaa gaacagaaag aaaggaaaac cacagaaatg caaagtccaa 1801 tgcagccagt ggaccaagcg agccttccag gtcactgcag ggaacctcca ccatgggaaa 1861 atgaagccac agagagaatt tatcatttcg tggtggggca gatggtttat tatcagtgcg 1921 tccagggata cagggctcta cacagaggtc ctgctgagag cgtctgcaaa atgacccacg 1981 ggaagacaag gtggacccag ccccagctca tatgcacagg tgaaatggag accagtcagt 2041 ttccaggtga agagaagcct caggcaagcc ccgaaggccg tcctgagagt gagacttcct 2101 gcctcgtcac aacagcagat tttcaaatac aggcagaaat ggctgcagcc atggagacgt 2161 ccatatttgc agcagagtac caggtagcag tggccggctg tgttttcctg ctgatcagcg 2221 tcctcctcct gagtgggctc acctggcagc ggagacagag gaagagtaga agaacaatct 2281 aggaattctg cagtcgacgg taccgcgggc ccgggatcca ccggatctag ataactgatc 2341 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc 2401 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct 2461 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca 2521 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaacgcgt aaattgtaag 2581 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca 2641 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag 2701 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg 2761 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt 2821 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag 2881 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc 2941 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc 3001 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac 3061 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc 3121 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg 3181 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg 3241 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca 3301 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact 3361 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta 3421 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag 3481 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg 3541 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga 3601 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt 3661 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct 3721 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg 3781 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt 3841 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc 3901 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc 3961 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga 4021 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag 4081 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat 4141 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg 4201 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc 4261 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta 4321 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg 4381 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc 4441 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg 4501 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa 4561 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt 4621 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc 4681 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt 4741 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac 4801 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag 4861 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg 4921 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc 4981 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag 5041 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt 5101 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac 5161 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc 5221 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt 5281 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt 5341 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc 5401 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt 5461 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca 5521 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt 5581 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt 5641 attaccgcca tgcat //