LOCUS Exported 6123 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Bacterial expression of mutant human alpha synuclein. ACCESSION . VERSION . KEYWORDS pET11a asyn Q99E (TAC-->TAT) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6123) TITLE Lashuel Lab plasmids REFERENCE 2 (bases 1 to 6123) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6123 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 193..225 /codon_start=1 /product="leader peptide from bacteriophage T7 gene 10" /label=T7 tag (gene 10 leader) /note="promotes efficient translation in E. coli" /translation="MASMTGGQQMG" primer_bind complement(303..321) /label=T7 Term /note="T7 terminator, reverse primer" terminator 317..364 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" promoter complement(539..567) /label=tet promoter /note="E. coli promoter for tetracycline efflux protein gene" primer_bind complement(594..612) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 680..784 /gene="bla" /label=AmpR promoter CDS 785..1645 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(1003..1022) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 1816..2403 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 2304..2323 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 2557..2574 /label=L4440 /note="L4440 vector, forward primer" misc_feature 2589..2728 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(2654..2673) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 2814..2836 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(2830..3021) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4333..5415) /codon_start=1 /gene="lacI" /product="lac repressor" /label=lacI /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" primer_bind 5377..5396 /label=LacI-R /note="LacI, reverse primer" promoter complement(5416..5493) /gene="lacI" /label=lacI promoter primer_bind 5699..5718 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind 5802..5821 /label=T7 /note="T7 promoter, forward primer" promoter 5802..5820 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 5821..5845 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." RBS 5860..5882 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" ORIGIN 1 cagaagacag tggagggagc ggggagcatt gcagcagcca ctggctttgt caaaaaggac 61 gagttgggca agaatgaaga aggagcccca caggaaggaa ttctggaaga tatgcctgtg 121 gatcctgaca atgaggctta tgaaatgcct tctgaggaag ggtatcaaga ctatgaacct 181 gaagcctaac atatggctag catgactggt ggacagcaaa tgggtcgcgg atcccgggta 241 cctcgagtcg accgatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc 301 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt 361 tttgctgaaa ggaggaacta tatccggata tcccgcaaga ggcccggcag taccggcata 421 accaagccta tgcctacagc atccagggtg acggtgccga ggatgacgat gagcgcattg 481 ttagatttca tacacggtgc ctgactgcgt tagcaattta actgtgataa actaccgcat 541 taaagcttat cgatgataag ctgtcaaaca tgagaattct tgaagacgaa agggcctcgt 601 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg 661 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa 721 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa 781 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct 841 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg 901 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg 961 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt 1021 atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga 1081 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga 1141 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac 1201 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg 1261 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac 1321 gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct 1381 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct 1441 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg 1501 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat 1561 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg 1621 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat 1681 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct 1741 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa 1801 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa 1861 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc 1921 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta 1981 gttaggccac cacttcaaga acctgtagca ccgcctacat acctcgctct gctaatcctg 2041 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga 2101 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc 2161 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc 2221 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga 2281 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt 2341 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg 2401 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac 2461 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga 2521 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg 2581 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat 2641 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct 2701 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc 2761 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag 2821 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag 2881 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc 2941 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc 3001 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt 3061 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc 3121 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag 3181 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg 3241 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg 3301 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc 3361 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta 3421 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg 3481 cacccgtggc caggacccaa cgctgcccga gatgcgccgc gtgcggctgc tggagatggc 3541 ggacgcgatg gatatgttct gccaagggtt ggtttgcgca ttcacagttc tccgcaagaa 3601 ttgattggct ccaattcttg gagtggtgaa tccgttagcg aggtgccgcc ggcttccatt 3661 caggtcgagg tggcccggct ccatgcaccg cgacgcaacg cggggaggca gacaaggtat 3721 agggcggcgc ctacaatcca tgccaacccg ttccatgtgc tcgccgaggc ggcataaatc 3781 gccgtgacga tcagcggtcc aatgatcgaa gttaggctgg taagagccgc gagcgatcct 3841 tgaagctgtc cctgatggtc gtcatctacc tgcctggaca gcatggcctg caacgcgggc 3901 atcccgatgc cgccggaagc gagaagaatc ataatgggga aggccatcca gcctcgcgtc 3961 gcgaacgcca gcaagacgta gcccagcgcg tcggccgcca tgccggcgat aatggcctgc 4021 ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag 4081 attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg 4141 ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca 4201 gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg 4261 aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta 4321 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa 4381 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct 4441 tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg 4501 cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa 4561 cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc 4621 accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt 4681 ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa 4741 accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt 4801 gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc 4861 taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc 4921 gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa 4981 cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata 5041 gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc 5101 ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg 5161 agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac 5221 gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt 5281 cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg 5341 gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa 5401 cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat 5461 accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg 5521 actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa 5581 ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca 5641 tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg 5701 tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga 5761 tgcgtccggc gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg 5821 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga 5881 gatatacata tggatgtatt catgaaagga ctttcaaagg ccaaggaggg agttgtggct 5941 gctgctgaga aaaccaaaca gggtgtggca gaagcagcag gaaagacaaa agagggtgtt 6001 ctctatgtag gctccaaaac caaggaggga gtggtgcatg gtgtggcaac agtggctgag 6061 aagaccaaag agcaagtgac aaatgttgga ggagcagtgg tgacgggtgt gacagcagta 6121 gcc //