LOCUS       sequence-202102-        6100 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expression of the human Huntingtin Exon1 protein containing 29Q in 
            E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6100)
  AUTHORS   Vieweg S, Ansaloni A, Wang ZM, Warner JB, Lashuel HA
  TITLE     An Intein-based Strategy for the Production of Tag-free Huntingtin 
            Exon 1 Proteins Enables New Insights into the Polyglutamine 
            Dependence of Httex1 Aggregation and Fibril Formation.
  JOURNAL   J Biol Chem. 2016 Jun 3;291(23):12074-86. doi: 
            10.1074/jbc.M116.713982. Epub 2016 Mar 21.
  PUBMED    27002149
REFERENCE   2  (bases 1 to 6100)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1074/jbc.M116.713982"; journalName: "J Biol Chem"; date: 
            "2016-06-3- 3"; volume: "291"; issue: "23"; pages: "12074-86"
COMMENT     Sequence Label: pTWIN1-His6-Ssp-Httex1-29Q
FEATURES             Location/Qualifiers
     source          1..6100
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      42..128
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     263..282
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        263..281
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    282..306
                     /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             321..343
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             354..371
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1189..1207)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1203..1250
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1310..1414
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1415..1483,1484..2275)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1633..1652)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2317..2830)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2512..2533)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2724..2743
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      2941..3529
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3430..3449
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3683..3700
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3722..3741)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3882..3904
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3898..4089)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4646..5728)
                     /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     5690..5709
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5729..5806)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      5959..6045
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gacgcccgcc ataaactgcc aggaattaat tccccaggca tcaaataaaa cgaaaggctc
       61 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
      121 ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg
      181 caggacgccc gccataaact gccaggaatt ggggatcgga attaattccc ggtttaaacc
      241 ggggatctcg atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac
      301 aattcccctc tagaaataat tttgtttaac tttaagaagg agatatacat atgcatcatc
      361 atcatcacca tagcggtgca attagcggtg atagcctgat tagcctggca agcaccggta
      421 aacgtgttag cattaaagat ctgctggatg agaaagattt tgagatttgg gccattaacg
      481 agcagaccat gaaactggaa agcgcaaaag ttagccgtgt tttttgcacc ggtaaaaaac
      541 tggtgtatat cctgaaaacc cgtctgggtc gtaccattaa agcaaccgca aatcatcgtt
      601 ttctgaccat tgatggttgg aaacgtctgg atgaactgag cctgaaagaa catattgcac
      661 tgcctcgtaa actggaaagt agcagcctgc aactgagtcc ggaaattgaa aaactgagcc
      721 agagcgatat ctattgggat agcattgtta gcattaccga aaccggtgtt gaagaagttt
      781 ttgatctgac cgttccgggt ccgcacaatt ttgttgcaaa tgatattatt gtgcacaacg
      841 ccaccctgga aaaactgatg aaagcatttg aaagcctgaa aagctttcag cagcagcaac
      901 aacagcagca acagcaacag cagcagcagc agcaacaaca gcagcagcaa cagcaacaac
      961 aacaacaaca gcagcctcct ccgcctccgc caccgcctcc acctccgcag ctgccgcagc
     1021 ctccacctca ggcacaaccg ctgctgcctc agccgcagcc acctcctccg ccaccaccgc
     1081 caccgcctgg tccagcagtt gcagaagaac cgctgcatcg tccgtaactg caggaagggg
     1141 atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat
     1201 aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag
     1261 gaactatatc cggataacta cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     1321 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     1381 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     1441 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     1501 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     1561 caacagcggt aagatccttg agagttttcg ccccgaagaa cgtttcccaa tgatgagcac
     1621 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact
     1681 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     1741 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     1801 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     1861 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     1921 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     1981 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     2041 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     2101 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     2161 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     2221 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     2281 agaccaagtt tactcatata tactttagat tgatttaccc cggttgataa tcagaaaagc
     2341 cccaaaaaca ggaagattgt ataagcaaat atttaaattg taaacgttaa tattttgtta
     2401 aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc
     2461 aaaatccctt ataaatcaaa agaatagccc gagatagggt tgagtgttgt tccagtttgg
     2521 aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat
     2581 cagggcgatg gcccactacg tgaaccatca cccaaatcaa gttttttggg gtcgaggtgc
     2641 cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag
     2701 ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg
     2761 gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta
     2821 cagggcgcgt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2881 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2941 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     3001 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     3061 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     3121 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     3181 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     3241 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     3301 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3361 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3421 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3481 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3541 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3601 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3661 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagctatgg tgcactctca
     3721 gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga
     3781 ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     3841 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     3901 gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg
     3961 gtcgtgcagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc
     4021 cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg
     4081 tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat
     4141 gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga
     4201 acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca
     4261 gggtcaatgc cagccgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc
     4321 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     4381 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     4441 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     4501 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     4561 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     4621 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4681 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     4741 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     4801 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     4861 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     4921 ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     4981 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     5041 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     5101 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     5161 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     5221 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     5281 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     5341 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     5401 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     5461 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     5521 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     5581 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     5641 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     5701 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     5761 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     5821 tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc
     5881 accgccgccg caaggaatgg tgcatgccgg catgccgccc tttcgtcttc aagaattaat
     5941 tcccaattcc ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt
     6001 tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat
     6061 ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
//
