LOCUS       sequence-222747-        6082 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of Htt fragment containing exon 1, with A2C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6082)
  AUTHORS   Warner JB 4th, Ruff KM, Tan PS, Lemke EA, Pappu RV, Lashuel HA
  TITLE     Monomeric Huntingtin Exon 1 Has Similar Overall Structural Features 
            for Wild-Type and Pathological Polyglutamine Lengths.
  JOURNAL   J Am Chem Soc. 2017 Oct 18;139(41):14456-14469. doi: 
            10.1021/jacs.7b06659. Epub 2017 Oct 9.
  PUBMED    28937758
REFERENCE   2  (bases 1 to 6082)
  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: 
            "10.1021/jacs.7b06659"; journalName: "J Am Chem Soc"; date: 
            "2017-10-18- 18"; volume: "139"; issue: "41"; pages: "14456-14469"
COMMENT     Sequence Label: pTWIN1-His6-Ssp-Httex1-23Q-A2C-A60C
FEATURES             Location/Qualifiers
     source          1..6082
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      32..118
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     253..272
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        253..271
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    272..296
                     /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             311..333
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             344..361
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1161..1179)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1175..1222
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1282..1386
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1387..1455,1456..2247)
                     /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(1605..1624)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2289..2802)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2484..2505)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2696..2715
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      2913..3501
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3402..3421
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3655..3672
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3694..3713)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3854..3876
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3870..4061)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4618..5700)
                     /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     5662..5681
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5701..5778)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      5931..6017
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ataaactgcc aggaattaat tccccaggca tcaaataaaa cgaaaggctc agtcgaaaga
       61 ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc
      121 gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg caggacgccc
      181 gccataaact gccaggaatt ggggatcgga attaattccc ggtttaaacc ggggatctcg
      241 atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac aattcccctc
      301 tagaaataat tttgtttaac tttaagaagg agatatacat atgcatcatc atcatcacca
      361 tagcggtgca attagcggtg atagcctgat tagcctggca agcaccggta aacgtgttag
      421 cattaaagat ctgctggatg agaaagattt tgagatttgg gccattaacg agcagaccat
      481 gaaactggaa agcgcaaaag ttagccgtgt tttttgcacc ggtaaaaaac tggtgtatat
      541 cctgaaaacc cgtctgggtc gtaccattaa agcaaccgca aatcatcgtt ttctgaccat
      601 tgatggttgg aaacgtctgg atgaactgag cctgaaagaa catattgcac tgcctcgtaa
      661 actggaaagt agcagcctgc aactgagtcc ggaaattgaa aaactgagcc agagcgatat
      721 ctattgggat agcattgtta gcattaccga aaccggtgtt gaagaagttt ttgatctgac
      781 cgttccgggt ccgcacaatt ttgttgcaaa tgatattatt gtgcacaact gcaccctgga
      841 aaaactgatg aaagcatttg aaagcctgaa aagctttcag caacaacagc agcaacagca
      901 acagcagcag cagcagcaac aacagcagca gcaacagcaa caacaaccgc ctccgcctcc
      961 gccacctcct cctccaccac agctgcctca gcctccgcct cagtgccagc cgctgctgcc
     1021 acagcctcag ccacctccac cgcctccacc tccacctccg ggtccggcag ttgcagaaga
     1081 accgctgcat cgtccgtaac tgcaggaagg ggatccggct gctaacaaag cccgaaagga
     1141 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa
     1201 acgggtcttg aggggttttt tgctgaaagg aggaactata tccggataac tacgtcaggt
     1261 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     1321 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     1381 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     1441 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     1501 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     1561 cgccccgaag aacgtttccc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     1621 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     1681 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     1741 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     1801 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     1861 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     1921 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     1981 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     2041 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     2101 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     2161 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     2221 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     2281 attgatttac cccggttgat aatcagaaaa gccccaaaaa caggaagatt gtataagcaa
     2341 atatttaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     2401 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaatagc
     2461 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     2521 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     2581 cacccaaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     2641 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     2701 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     2761 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtaaaaggat ctaggtgaag
     2821 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     2881 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     2941 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3001 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     3061 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3121 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3181 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3241 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3301 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3361 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     3421 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3481 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3541 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     3601 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     3661 tcagtgagcg aggaagctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     3721 aagccagtat acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg
     3781 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     3841 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     3901 gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgca gcgattcaca gatgtctgcc
     3961 tgttcatccg cgtccagctc gttgagtttc tccagaagcg ttaatgtctg gcttctgata
     4021 aagcgggcca tgttaagggc ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg
     4081 ggatttctgt tcatgggggt aatgataccg atgaaacgag agaggatgct cacgatacgg
     4141 gttactgatg atgaacatgc ccggttactg gaacgttgtg agggtaaaca actggcggta
     4201 tggatgcggc gggaccagag aaaaatcact cagggtcaat gccagccgaa cgccagcaag
     4261 acgtagccca gcgcgtcggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt
     4321 ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca
     4381 agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag
     4441 agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg
     4501 acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc
     4561 atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca
     4621 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     4681 gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac
     4741 gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac
     4801 gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca
     4861 tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc
     4921 ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc
     4981 agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact
     5041 ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca
     5101 gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg
     5161 ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa
     5221 aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt
     5281 gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc
     5341 actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg
     5401 ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc
     5461 gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga
     5521 ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc
     5581 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
     5641 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
     5701 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
     5761 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
     5821 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgcc
     5881 ggcatgccgc cctttcgtct tcaagaatta attcccaatt ccccaggcat caaataaaac
     5941 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     6001 tcctgagtag gacaaatccg ccgggagcgg atttgaacgt tgcgaagcaa cggcccggag
     6061 ggtggcgggc aggacgcccg cc
//
