LOCUS Exported 5085 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Express a EGFP-human MAP1-LC3 in mammalian cells. ACCESSION . VERSION . KEYWORDS pEGFP-hLC3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5085) AUTHORS Tanida I, Tanida-Miyake E, Ueno T, Kominami E TITLE The human homolog of Saccharomyces cerevisiae Apg7p is a Protein-activating enzyme for multiple substrates including human Apg12p, GATE-16, GABARAP, and MAP-LC3. JOURNAL J Biol Chem. 2001 Jan 19;276(3):1701-6. Epub 2000 Nov 28. PUBMED 11096062 REFERENCE 2 (bases 1 to 5085) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5085 /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" regulatory 243..252 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 249..965 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 249..965 /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(294..315) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(555..574) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 902..923 /label=EGFP-C /note="EGFP, forward primer" CDS 981..1355 /codon_start=1 /gene="MAP1LC3B" /product="human LC3 protein" /label=LC3B /note="Atg8 family member found on autophagosomes" /translation="MPSEKTFKQRRTFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLP VLDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESEKD EDGFLYMVYASQETFGMKLSV" polyA_signal 1509..1630 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1546..1565) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1600..1619 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(1637..2092) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1774..1795) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 1986..2005 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2119..2223 /gene="bla" /label=AmpR promoter promoter 2225..2582 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2248..2268) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 2433..2568 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2495..2514 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 2617..3411 /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(2671..2690) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3281..3300 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(3599..3618) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 3643..3690 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4019..4607 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4508..4527 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 4782..5085 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" ORIGIN 1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt 61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac 121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg 181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg 241 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 301 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 361 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 421 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 481 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 541 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 601 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 661 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 721 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 781 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 841 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 901 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 961 acaagtccgg actcagatct atgccgtcgg agaagacctt caagcagcgc cgcaccttcg 1021 aacaaagagt agaagatgtc cgacttattc gagagcagca tccaaccaaa atcccggtga 1081 taatagaacg atacaagggt gagaagcagc ttcctgttct ggataaaaca aagttccttg 1141 tacctgacca tgtcaacatg agtgagctca tcaagataat tagaaggcgc ttacagctca 1201 atgctaatca ggccttcttc ctgttggtga acggacacag catggtcagc gtctccacac 1261 caatctcaga ggtgtatgag agtgagaaag atgaagatgg attcctgtac atggtctacg 1321 cctcccagga gacgttcggg atgaaattgt cagtgtaagt cgacggtacc gcgggcccgg 1381 gatccaccgg atctagataa ctgatcataa tcagccatac cacatttgta gaggttttac 1441 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg 1501 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa 1561 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca 1621 atgtatctta acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt 1681 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca 1741 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta 1801 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta 1861 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg 1921 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga 1981 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg 2041 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc 2101 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat 2161 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag 2221 agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc 2281 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt 2341 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt 2401 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg 2461 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct 2521 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca 2581 aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac 2641 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca 2701 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt 2761 gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg 2821 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga 2881 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct 2941 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg 3001 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg 3061 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc 3121 gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat 3181 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac 3241 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt 3301 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct 3361 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc 3421 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca 3481 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga 3541 tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg 3601 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga 3661 ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc 3721 actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt 3781 ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg 3841 caggccctgc catagcctca ggttactcat atatacttta gattgattta aaacttcatt 3901 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt 3961 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt 4021 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag 4081 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca 4141 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca 4201 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg 4261 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg 4321 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct 4381 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga 4441 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc 4501 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg 4561 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg 4621 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt 4681 tatcccctga ttctgtggat aaccgtatta ccgccatgca ttagttatta atagtaatca 4741 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta 4801 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat 4861 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg 4921 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac 4981 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt 5041 cctacttggc agtacatcta cgtattagtc atcgctatta ccatg //