LOCUS       Exported                6119 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  screen AB42 variants for aggregation.
ACCESSION   .
VERSION     .
KEYWORDS    AB-GFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6119)
  AUTHORS   Wurth C, Guimard NK, Hecht MH
  TITLE     Mutations that reduce aggregation of the Alzheimer's Abeta42 
            peptide: an unbiased search for the sequence determinants of Abeta 
            amyloidogenesis.
  JOURNAL   J Mol Biol. 2002 Jun 21;319(5):1279-90.
  PUBMED    12079364
REFERENCE   2  (bases 1 to 6119)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6119
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     terminator      26..73
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(173..883)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(213..233)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     828..856
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     RBS             1056..1078
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1093..1117
                     /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)."
     primer_bind     complement(1117..1136)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1118..1136)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1220..1239)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1445..1522
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1523..2605
                     /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     complement(1542..1561)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3414..3605
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3599..3621)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3707..3849
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3762..3781
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3864..3881)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4035..4623)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4115..4134)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4734..4753)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4745..5560
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4818..4837)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5653..6108)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5790..5811)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6002..6021
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagggtacct tatttgtaga
      181 gctcatccat gccatgtgta atcccagcag cagttacaaa ctcaagaagg accatgtggt
      241 cacgcttttc gttgggatct ttcgaaaggg cagattgtgt cgacaggtaa tggttgtctg
      301 gtaaaaggac agggccatcg ccaattggag tattttgttg ataatggtct gctagttgaa
      361 cggaaccatc ttcaatgttg tggcgaattt tgaagttagc tttgattcca ttcttttgtt
      421 tgtctgccgt gatgtataca ttgtgtgagt tatagttgta ctcgagtttg tgtccgagaa
      481 tgtttccatc ttctttaaaa tcaataccct ttaactcgat acgattaaca agggtatcac
      541 cttcaaactt gacttcagca cgcgtcttgt agttcccgtc atctttgaaa gatatagtgc
      601 gttcctgtac ataaccttcg ggcatggcac tcttgaaaaa gtcatgccgt ttcatgtgat
      661 ccggataacg ggaaaagcat tgaacaccat aggtcagagt agtgacaagt gttggccatg
      721 gaacaggtag ttttccagta gtgcaaataa atttaagggt gagttttccg tatgtagcat
      781 caccttcacc ctctccactg acagaaaatt tgtgcccatt aacatcacca tctaattcaa
      841 caagaattgg gacaactcca gtgaaaagtt cttctccttt gctgaattcg ccagaaccag
      901 cagcggagcc agcggatccc gcaatcacca cgccgcccac catcaggcca ataatcgcgc
      961 ctttgttaga gcccacatct tccgcaaaaa acaccagttt ctgatgatgc acttcatagc
     1021 cagaatcatg gcgaaattcc gcatccatat gtatatctcc ttcttaaagt taaacaaaat
     1081 tatttctaga ggggaattgt tatccgctca caattcccct atagtgagtc gtattaattt
     1141 cgcgggatcg agatctcgat cctctacgcc ggacgcatcg tggccggcat caccggcgcc
     1201 acaggtgcgg ttgctggcgc ctatatcgcc gacatcaccg atggggaaga tcgggctcgc
     1261 cacttcgggc tcatgagcgc ttgtttcggc gtgggtatgg tggcaggccc cgtggccggg
     1321 ggactgttgg gcgccatctc cttgcatgca ccattccttg cggcggcggt gctcaacggc
     1381 ctcaacctac tactgggctg cttcctaatg caggagtcgc ataagggaga gcgtcgagat
     1441 cccggacacc atcgaatggc gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga
     1501 gagtcaattc agggtggtga atgtgaaacc agtaacgtta tacgatgtcg cagagtatgc
     1561 cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa
     1621 aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc
     1681 acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct
     1741 gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag
     1801 cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa
     1861 tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc
     1921 cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca
     1981 gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg gcgtggagca
     2041 tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttctgtctc
     2101 ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa ttcagccgat
     2161 agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca tgcaaatgct
     2221 gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc
     2281 aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg tagtgggata
     2341 cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca aacaggattt
     2401 tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt
     2461 gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc tggcgcccaa
     2521 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     2581 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtaagttag ctcactcatt
     2641 aggcaccggg atctcgaccg atgcccttga gagccttcaa cccagtcagc tccttccggt
     2701 gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc atgcaactcg
     2761 taggacaggt gccggcagcg ctctgggtca ttttcggcga ggaccgcttt cgctggagcg
     2821 cgacgatgat cggcctgtcg cttgcggtat tcggaatctt gcacgccctc gctcaagcct
     2881 tcgtcactgg tcccgccacc aaacgtttcg gcgagaagca ggccattatc gccggcatgg
     2941 cggccccacg ggtgcgcatg atcgtgctcc tgtcgttgag gacccggcta ggctggcggg
     3001 gttgccttac tggttagcag aatgaatcac cgatacgcga gcgaacgtga agcgactgct
     3061 gctgcaaaac gtctgcgacc tgagcaacaa catgaatggt cttcggtttc cgtgtttcgt
     3121 aaagtctgga aacgcggaag tcagcgccct gcaccattat gttccggatc tgcatcgcag
     3181 gatgctgctg gctaccctgt ggaacaccta catctgtatt aacgaagcgc tggcattgac
     3241 cctgagtgat ttttctctgg tcccgccgca tccataccgc cagttgttta ccctcacaac
     3301 gttccagtaa ccgggcatgt tcatcatcag taacccgtat cgtgagcatc ctctctcgtt
     3361 tcatcggtat cattaccccc atgaacagaa atccccctta cacggaggca tcagtgacca
     3421 aacaggaaaa aaccgccctt aacatggccc gctttatcag aagccagaca ttaacgcttc
     3481 tggagaaact caacgagctg gacgcggatg aacaggcaga catctgtgaa tcgcttcacg
     3541 accacgctga tgagctttac cgcagctgcc tcgcgcgttt cggtgatgac ggtgaaaacc
     3601 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     3661 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc
     3721 agtcacgtag cgatagcgga gtgtatactg gcttaactat gcggcatcag agcagattgt
     3781 actgagagtg caccatatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     3841 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     3901 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     3961 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4021 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4081 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4141 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4201 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4261 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4321 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4381 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4441 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     4501 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     4561 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     4621 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     4681 taagggattt tggtcatgaa caataaaact gtctgcttac ataaacagta atacaagggg
     4741 tgttatgagc catattcaac gggaaacgtc ttgctctagg ccgcgattaa attccaacat
     4801 ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac
     4861 aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg
     4921 tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat
     4981 gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac
     5041 tgcgatcccc gggaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa
     5101 tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg
     5161 tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa tgaataacgg
     5221 tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg
     5281 gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc atggtgattt
     5341 ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg
     5401 agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt
     5461 ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgaa
     5521 taaattgcag tttcatttga tgctcgatga gtttttctaa gaattaattc atgagcggat
     5581 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     5641 aagtgccacc tgaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt
     5701 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     5761 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     5821 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     5881 aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     5941 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     6001 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     6061 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgcca
//
