LOCUS Exported 5668 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression of Rluc-LC3G120A. Can be used for measuring autophagic flux.. ACCESSION . VERSION . KEYWORDS pRluc-LC3G120A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5668) AUTHORS Farkas T, Jaattela M TITLE Renilla Luciferase-LC3 Based Reporter Assay for Measuring Autophagic Flux. JOURNAL Methods Enzymol. 2017;588:1-13. doi: 10.1016/bs.mie.2016.09.072. Epub 2016 Nov 12. PUBMED 28237095 REFERENCE 2 (bases 1 to 5668) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5668 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..304 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 305..508 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 459..479 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" CDS 553..1485 /codon_start=1 /product="luciferase from the anthozoan coelenterate Renilla reniformis (sea pansy)" /label=Rluc /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW FELLNLPKKIIFVGHDWGAALAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ" CDS 1504..1926 /codon_start=1 /gene="Map1lc3b" /product="rat LC3 protein" /label=LC3 /note="Atg8 family member found on autophagosomes" /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV LDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE DGFLYMVYASQETFA*ALAVTYMSALKATATGREPCL" polyA_signal 2396..2517 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2433..2452) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2487..2506 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2524..2979) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2661..2682) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2873..2892 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3006..3110 /gene="bla" /label=AmpR promoter promoter 3112..3469 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3135..3155) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3320..3455 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3382..3401 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3504..4298 /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(3558..3577) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4168..4187 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4486..4505) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4530..4577 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4906..5494 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5395..5414 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta 541 ccggtcgcca ccatgacttc gaaagtttat gatccagaac aaaggaaacg gatgataact 601 ggtccgcagt ggtgggccag atgtaaacaa atgaatgttc ttgattcatt tattaattat 661 tatgattcag aaaaacatgc agaaaatgct gttatttttt tacatggtaa cgcggcctct 721 tcttatttat ggcgacatgt tgtgccacat attgagccag tagcgcggtg tattatacca 781 gaccttattg gtatgggcaa atcaggcaaa tctggtaatg gttcttatag gttacttgat 841 cattacaaat atcttactgc atggtttgaa cttcttaatt taccaaagaa gatcattttt 901 gtcggccatg attggggtgc tgctttggca tttcattata gctatgagca tcaagataag 961 atcaaagcaa tagttcacgc tgaaagtgta gtagatgtga ttgaatcatg ggatgaatgg 1021 cctgatattg aagaagatat tgcgttgatc aaatctgaag aaggagaaaa aatggttttg 1081 gagaataact tcttcgtgga aaccatgttg ccatcaaaaa tcatgagaaa gttagaacca 1141 gaagaatttg cagcatatct tgaaccattc aaagagaaag gtgaagttcg tcgtccaaca 1201 ttatcatggc ctcgtgaaat cccgttagta aaaggtggta aacctgacgt tgtacaaatt 1261 gttaggaatt ataatgctta tctacgtgca agtgatgatt taccaaaaat gtttattgaa 1321 tcggacccag gattcttttc caatgctatt gttgaaggtg ccaagaagtt tcctaatact 1381 gaatttgtca aagtaaaagg tcttcatttt tcgcaagaag atgcacctga tgaaatggga 1441 aaatatatca aatcgttcgt tgagcgagtt ctcaaaaatg aacaatccgg actcagatct 1501 agaccgtccg agaagacctt caaacagcgc cggagcttcg aacaaagagt ggaagatgtc 1561 cggctcatcc gggagcagca ccccaccaag atcccagtga ttatagagcg atacaagggt 1621 gagaagcagc tgcccgtcct ggacaagacc aagttccttg tacctgatca cgtgaatatg 1681 agcgaactca tcaagataat tagaaggcgc ctgcagctca atgctaacca agccttcttc 1741 ctcctggtga atgggcacag catggtgagt gtgtccacac ccatctctga agtgtacgag 1801 agcgagagag atgaagacgg cttcctgtac atggtctatg cctcccagga gacgttcgcg 1861 taagcactgg ctgttacata catgtcagct ctgaaggcaa cagcaacagg aagagagcca 1921 tgcttgtgac agacatacag ccacttccaa ctaaagcaag cctctgctcc ctgctacctg 1981 catggagccc actgtgacac tcagaccatc cccggtcact cactcactcg tgtctgagaa 2041 tctcagtgag agctgcctct gtcacggacg gaagccaaca cagccacctc tcgacctggc 2101 tccccacagc acccaccctc cctgcatgca gctgtccctg ctaaccccca atgttatgtt 2161 acactgtgta agtccccact gctgccgtgt gtgggttgtg tacgtcgtca tgtccctggt 2221 ttataactat ggtgcgaatt ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc 2281 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa 2341 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt 2401 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa 2461 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg 2521 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct 2581 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg 2641 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact 2701 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac 2761 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga 2821 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga 2881 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca 2941 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa 3001 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca 3061 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg 3121 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc 3181 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc 3241 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt 3301 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc 3361 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct 3421 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag 3481 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg 3541 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg 3601 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc 3661 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga 3721 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc 3781 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag 3841 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat 3901 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg 3961 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca 4021 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct 4081 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg 4141 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg 4201 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc 4261 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat 4321 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta 4381 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg 4441 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga 4501 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg 4561 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac 4621 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc 4681 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat 4741 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4801 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc 4861 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt 4921 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt 4981 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5041 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5101 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5161 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5221 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5281 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5341 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5401 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5461 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5521 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5581 tgtggataac cgtattaccg ccatgcatta gttattaata gtaatcaatt acggggtcat 5641 tagttcatag cccatatatg gagttccg //