LOCUS       Exported                6124 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression of mutant human alpha synuclein.
ACCESSION   .
VERSION     .
KEYWORDS    pET11a asyn Q79N Q109N (TAC-->TAT)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6124)
  TITLE     Lashuel Lab plasmids
REFERENCE   2  (bases 1 to 6124)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6124
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      28..616
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     517..536
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     770..787
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    802..941
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(867..886)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1027..1049
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1043..1234)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2546..3628)
                     /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     3590..3609
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3629..3706)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3912..3931
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4015..4034
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4015..4033
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4034..4058
                     /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             4073..4095
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4529..4561
                     /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(4639..4657)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4653..4700
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(4875..4903)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(4930..4948)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5016..5120
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5121..5981
                     /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(5339..5358)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
       61 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
      121 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
      181 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
      241 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
      301 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
      361 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
      421 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
      481 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
      541 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
      601 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
      661 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
      721 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
      781 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
      841 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
      901 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
      961 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     1021 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     1081 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     1141 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     1201 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     1261 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     1321 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     1381 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     1441 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     1501 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     1561 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     1621 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     1681 gcacgatcat gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc
     1741 tgctggagat ggcggacgcg atggatatgt tctgccaagg gttggtttgc gcattcacag
     1801 ttctccgcaa gaattgattg gctccaattc ttggagtggt gaatccgtta gcgaggtgcc
     1861 gccggcttcc attcaggtcg aggtggcccg gctccatgca ccgcgacgca acgcggggag
     1921 gcagacaagg tatagggcgg cgcctacaat ccatgccaac ccgttccatg tgctcgccga
     1981 ggcggcataa atcgccgtga cgatcagcgg tccaatgatc gaagttaggc tggtaagagc
     2041 cgcgagcgat ccttgaagct gtccctgatg gtcgtcatct acctgcctgg acagcatggc
     2101 ctgcaacgcg ggcatcccga tgccgccgga agcgagaaga atcataatgg ggaaggccat
     2161 ccagcctcgc gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc
     2221 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     2281 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     2341 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     2401 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     2461 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     2521 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     2581 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     2641 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     2701 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     2761 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     2821 ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     2881 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     2941 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     3001 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     3061 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     3121 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     3181 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     3241 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     3301 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     3361 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     3421 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     3481 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     3541 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     3601 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     3661 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     3721 tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc
     3781 accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg
     3841 gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga
     3901 tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg
     3961 atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg aaattaatac
     4021 gactcactat aggggaattg tgagcggata acaattcccc tctagaaata attttgttta
     4081 actttaagaa ggagatatac atatggatgt attcatgaaa ggactttcaa aggccaagga
     4141 gggagttgtg gctgctgctg agaaaaccaa acagggtgtg gcagaagcag caggaaagac
     4201 aaaagagggt gttctctatg taggctccaa aaccaaggag ggagtggtgc atggtgtggc
     4261 aacagtggct gagaagacca aagagcaagt gacaaatgtt ggaggagcag tggtgacggg
     4321 tgtgacagca gtagccaaca agacagtgga gggagcgggg agcattgcag cagccactgg
     4381 ctttgtcaaa aaggaccagt tgggcaagaa tgaagaagga gccccaaacg aaggaattct
     4441 ggaagatatg cctgtggatc ctgacaatga ggcttatgaa atgccttctg aggaagggta
     4501 tcaagactat gaacctgaag cctaacatat ggctagcatg actggtggac agcaaatggg
     4561 tcgcggatcc cgggtacctc gagtcgaccg atccggctgc taacaaagcc cgaaaggaag
     4621 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     4681 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatatccc gcaagaggcc
     4741 cggcagtacc ggcataacca agcctatgcc tacagcatcc agggtgacgg tgccgaggat
     4801 gacgatgagc gcattgttag atttcataca cggtgcctga ctgcgttagc aatttaactg
     4861 tgataaacta ccgcattaaa gcttatcgat gataagctgt caaacatgag aattcttgaa
     4921 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     4981 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5041 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5101 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     5161 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5221 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5281 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     5341 tatgtggcgc ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac
     5401 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     5461 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     5521 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     5581 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     5641 acgagcgtga caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg
     5701 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     5761 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     5821 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     5881 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     5941 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     6001 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6061 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6121 caga
//
