LOCUS Exported 6848 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION evaluate autophagy. ACCESSION . VERSION . KEYWORDS AV0-BWS-2.0-mC-G-CL3B SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6848) TITLE Oka Lab plasmids REFERENCE 2 (bases 1 to 6848) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6848 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 75..1253 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 306..1244 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1201..1221 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" regulatory 1280..1289 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1286..1993 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(1431..1449) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1699..1718 /label=mCherry-F /note="mCherry, forward primer" protein_bind 2021..2045 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" regulatory 2054..2063 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2060..2776 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 2060..2776 /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(2105..2126) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2366..2385) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2713..2734 /label=EGFP-C /note="EGFP, forward primer" CDS 2804..3178 /codon_start=1 /gene="MAP1LC3B" /product="human LC3 protein" /label=LC3B /note="Atg8 family member found on autophagosomes" /translation="MPSEKTFKQRRTFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLP VLDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESEKD EDGFLYMVYASQETFGMKLSV" misc_feature 3200..3788 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3253..3273) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3671..3682) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3809..3890 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3846..3865) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 3900..3919 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" repeat_region 3943..4083 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3943..4072 /label=AAV2 ITR rep_origin 4158..4613 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4245..4264) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4455..4476 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4630..4649) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4749..4771 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4809..4827) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4895..4999 /gene="bla" /label=AmpR promoter CDS 5000..5860 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(5218..5237) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6031..6619 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6520..6539 /label=pBR322ori-F /note="pBR322 origin, forward primer" repeat_region 6681..6810 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" ORIGIN 1 gatggataaa gttttaaaca gagaggaatc tttgcagcta atggaccttc taggtcttga 61 aaggagtggg aattggctcc ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc 121 ccgagaagtt ggggggaggg gtcggcaatt gaaccggtgc ctagagaagg tggcgcgggg 181 taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt gggggagaac 241 cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt gccgccagaa 301 cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct ctttacgggt tatggccctt 361 gcgtgccttg aattacttcc acctggctgc agtacgtgat tcttgatccc gagcttcggg 421 ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg agccccttcg cctcgtgctt 481 gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg aatctggtgg caccttcgcg 541 cctgtctcgc tgctttcgat aagtctctag ccatttaaaa tttttgatga cctgctgcga 601 cgcttttttt ctggcaagat agtcttgtaa atgcgggcca agatctgcac actggtattt 661 cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca tgttcggcga 721 ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa gctggccggc 781 ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg gcaaggctgg 841 cccggtcggc accagttgcg tgagcggaaa gatggccgct tcccggccct gctgcaggga 901 gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc acacaaagga 961 aaagggcctt tccgtcctca gccgtcgctt catgtgactc cacggagtac cgggcgccgt 1021 ccaggcacct cgattagttc tcgagctttt ggagtacgtc gtctttaggt tggggggagg 1081 ggttttatgc gatggagttt ccccacactg agtgggtgga gactgaagtt aggccagctt 1141 ggcacttgat gtaattctcc ttggaatttg ccctttttga gtttggatct tggttcattc 1201 tcaagcctca gacagtggtt caaagttttt ttcttccatt tcaggtgtcg tgaggaattg 1261 gatccgtttg gccggatctg ccaccatggt gagcaagggc gaggaggata acatggccat 1321 catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt 1381 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa 1441 ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta 1501 cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt 1561 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt 1621 gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac 1681 caacttcccc tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc 1741 cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct 1801 gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt 1861 gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga 1921 ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga 1981 cgagctgtac aagtccggac tcagatccgc tggcccagga acaagtttgt acaaaaaagc 2041 aggctttccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc 2101 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg 2161 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct 2221 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg 2281 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt 2341 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa 2401 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga 2461 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat 2521 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga 2581 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt 2641 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga 2701 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat 2761 ggacgagctg tacaagtccg gactcagatc cggtaccgaa ttcatgccgt cggagaagac 2821 cttcaagcag cgccgcacct tcgaacaaag agtagaagat gtccgactta ttcgagagca 2881 gcatccaacc aaaatcccgg tgataataga acgatacaag ggtgagaagc agcttcctgt 2941 tctggataaa acaaagttcc ttgtacctga ccatgtcaac atgagtgagc tcatcaagat 3001 aattagaagg cgcttacagc tcaatgctaa tcaggccttc ttcctgttgg tgaacggaca 3061 cagcatggtc agcgtctcca caccaatctc agaggtgtat gagagtgaga aagatgaaga 3121 tggattcctg tacatggtct atgcctccca ggagacgttc gggatgaaat tgtcagtgta 3181 agtcgacaat tctatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg 3241 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta 3301 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct 3361 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt 3421 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac 3481 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg 3541 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc 3601 gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg 3661 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct 3721 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc 3781 ctccccgcat cgataagtaa gtgactagaa cttgtttatt gcagcttata atggttacaa 3841 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg 3901 tggtttgtcc aaactcatct gcacgcgtgg cgcgccggcc gcaggaaccc ctagtgatgg 3961 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg 4021 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc 4081 aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac 4141 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt 4201 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt 4261 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc 4321 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga 4381 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc 4441 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg 4501 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct 4561 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg 4621 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac 4681 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt 4741 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag 4801 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc 4861 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt 4921 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata 4981 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt 5041 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc 5101 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat 5161 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct 5221 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca 5281 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg 5341 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa 5401 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg 5461 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga 5521 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg 5581 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt 5641 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg 5701 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc 5761 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca 5821 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc 5881 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat 5941 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc 6001 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg 6061 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct 6121 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct 6181 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct 6241 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg 6301 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc 6361 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga 6421 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg 6481 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta 6541 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg 6601 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg 6661 ctggcctttt gctcacatgt cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc 6721 ccgggcgtcg ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg 6781 gagtggccaa ctccatcact aggggttcct gcggccggcg cgccctgcag tttcccaagc 6841 tttgcaaa //