LOCUS Exported 5452 bp ds-DNA circular SYN 18-MAY-2021 DEFINITION Mammalian expression of rat LC3 fused to EGFP. ACCESSION . VERSION . KEYWORDS pEGFP-LC3-K51A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5452) AUTHORS Puri C, Manni MM, Vicinanza M, Hilcenko C, Zhu Y, Runwal G, Stamatakou E, Menzies FM, Mamchaoui K, Bitoun M, Rubinsztein DC TITLE A DNM2 Centronuclear Myopathy Mutation Reveals a Link between Recycling Endosome Scission and Autophagy. JOURNAL Dev Cell. 2020 Apr 20;53(2):154-168.e6. doi: 10.1016/j.devcel.2020.03.018. PUBMED 32315611 REFERENCE 2 (bases 1 to 5452) AUTHORS . TITLE Direct Submission JOURNAL Exported May 18, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5452 /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 984..1406 /codon_start=1 /gene="Map1lc3b" /product="rat LC3 protein" /label=LC3 /note="Atg8 family member found on autophagosomes" /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV LDKTAFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE DGFLYMVYASQETFGTALAVTYMSALKATATGREPCL" polyA_signal 1876..1997 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1913..1932) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1967..1986 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2004..2459) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2141..2162) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2353..2372 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2486..2590 /gene="bla" /label=AmpR promoter promoter 2592..2949 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2615..2635) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 2800..2935 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2862..2881 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 2984..3778 /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(3038..3057) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3648..3667 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(3966..3985) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4010..4057 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4386..4974 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4875..4894 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 5149..5452 /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 agaccgtccg agaagacctt caaacagcgc cggagcttcg 1021 aacaaagagt ggaagatgtc cggctcatcc gggagcagca ccccaccaag atcccagtga 1081 ttatagagcg atacaagggt gagaagcagc tgcccgtcct ggacaagacc gcgttccttg 1141 tacctgatca cgtgaatatg agcgaactca tcaagataat tagaaggcgc ctgcagctca 1201 atgctaacca agccttcttc ctcctggtga atgggcacag catggtgagt gtgtccacac 1261 ccatctctga agtgtacgag agcgagagag atgaagacgg cttcctgtac atggtctatg 1321 cctcccagga gacgttcggg acagcactgg ctgttacata catgtcagct ctgaaggcaa 1381 cagcaacagg aagagagcca tgcttgtgac agacatacag ccacttccaa ctaaagcaag 1441 cctctgctcc ctgctacctg catggagccc actgtgacac tcagaccatc cccggtcact 1501 cactcactcg tgtctgagaa tctcagtgag agctgcctct gtcacggacg gaagccaaca 1561 cagccacctc tcgacctggc tccccacagc acccaccctc cctgcatgca gctgtccctg 1621 ctaaccccca atgttatgtt acactgtgta agtccccact gctgccgtgt gtgggttgtg 1681 tacgtcgtca tgtccctggt ttataactat ggtgcgaatt ctgcagtcga cggtaccgcg 1741 ggcccgggat ccaccggatc tagataactg atcataatca gccataccac atttgtagag 1801 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat 1861 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc 1921 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa 1981 ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt 2041 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta 2101 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc 2161 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg 2221 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact 2281 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt 2341 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc 2401 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc 2461 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca 2521 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa 2581 aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga 2641 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 2701 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc 2761 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc 2821 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag 2881 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc 2941 ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg 3001 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca 3061 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt 3121 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg 3181 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga 3241 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca 3301 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct 3361 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac 3421 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc 3481 gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt 3541 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt 3601 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg 3661 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat 3721 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc 3781 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc 3841 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc 3901 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg 3961 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa 4021 agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag 4081 ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct 4141 tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc 4201 ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa 4261 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa 4321 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga 4381 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg 4441 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact 4501 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac 4561 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg 4621 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg 4681 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga 4741 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc 4801 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg 4861 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc 4921 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc 4981 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt 5041 cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcatta gttattaata 5101 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact 5161 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat 5221 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta 5281 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc 5341 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg 5401 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tg //