LOCUS       sequence-222653-        6061 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of Htt fragment containing amino acids 2-90, 
            with A2C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6061)
  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 6061)
  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-Htt2-90(A2C)-15Q-A60C
FEATURES             Location/Qualifiers
     source          1..6061
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      44..130
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     265..284
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        265..283
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    284..308
                     /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             323..345
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             356..373
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1152..1170)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1166..1213
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1273..1377
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1378..1446,1447..2238)
                     /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(1596..1615)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2280..2793)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2475..2496)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2687..2706
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      2904..3492
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3393..3412
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3646..3663
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3685..3704)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3845..3867
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3861..4052)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4609..5691)
                     /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     5653..5672
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5692..5769)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      5922..6008
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 aggacgcccg ccataaactg ccaggaatta attccccagg catcaaataa aacgaaaggc
       61 tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag
      121 taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg
      181 ggcaggacgc ccgccataaa ctgccaggaa ttggggatcg gaattaattc ccggtttaaa
      241 ccggggatct cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata
      301 acaattcccc tctagaaata attttgttta actttaagaa ggagatatac atatgcatca
      361 tcatcatcac catagcggtg caattagcgg tgatagcctg attagcctgg caagcaccgg
      421 taaacgtgtt agcattaaag atctgctgga tgagaaagat tttgagattt gggccattaa
      481 cgagcagacc atgaaactgg aaagcgcaaa agttagccgt gttttttgca ccggtaaaaa
      541 actggtgtat atcctgaaaa cccgtctggg tcgtaccatt aaagcaaccg caaatcatcg
      601 ttttctgacc attgatggtt ggaaacgtct ggatgaactg agcctgaaag aacatattgc
      661 actgcctcgt aaactggaaa gtagcagcct gcaactgagt ccggaaattg aaaaactgag
      721 ccagagcgat atctattggg atagcattgt tagcattacc gaaaccggtg ttgaagaagt
      781 ttttgatctg accgttccgg gtccgcacaa ttttgttgca aatgatatta ttgttcacaa
      841 ctgcaccctg gaaaaactga tgaaagcatt tgaaagcctg aaaagctttc agcagcagca
      901 acaacagcag caacagcaac aacaacaaca gcagcctccg cctccgcctc ctccaccacc
      961 tcctccacag ctgcctcagc ctccgcctca gtgccagccg ctgctgccac agcctcagcc
     1021 acctccaccg ccaccgcctc cacctccggg tccagcagtt gcagaagaac cgctgcatcg
     1081 tccgtaataa ctgcaggaag gggatccggc tgctaacaaa gcccgaaagg aagctgagtt
     1141 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     1201 gaggggtttt ttgctgaaag gaggaactat atccggataa ctacgtcagg tggcactttt
     1261 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     1321 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     1381 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     1441 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     1501 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     1561 gaacgtttcc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     1621 gttgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     1681 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     1741 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     1801 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     1861 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     1921 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     1981 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     2041 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     2101 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     2161 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     2221 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     2281 ccccggttga taatcagaaa agccccaaaa acaggaagat tgtataagca aatatttaaa
     2341 ttgtaaacgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     2401 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag cccgagatag
     2461 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     2521 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcacccaaat
     2581 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     2641 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     2701 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     2761 ccgccgcgct taatgcgccg ctacagggcg cgtaaaagga tctaggtgaa gatccttttt
     2821 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     2881 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     2941 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     3001 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     3061 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     3121 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     3181 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     3241 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     3301 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     3361 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     3421 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     3481 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     3541 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     3601 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     3661 gaggaagcta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta
     3721 tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc
     3781 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     3841 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
     3901 ctgcggtaaa gctcatcagc gtggtcgtgc agcgattcac agatgtctgc ctgttcatcc
     3961 gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc
     4021 atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg
     4081 ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat
     4141 gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg
     4201 cgggaccaga gaaaaatcac tcagggtcaa tgccagccga acgccagcaa gacgtagccc
     4261 agcgcgtcgg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg
     4321 ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg
     4381 ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc
     4441 ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc
     4501 atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga
     4561 gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
     4621 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4681 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
     4741 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
     4801 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
     4861 ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt
     4921 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
     4981 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
     5041 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
     5101 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
     5161 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
     5221 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
     5281 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
     5341 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
     5401 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
     5461 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
     5521 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     5581 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     5641 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     5701 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     5761 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     5821 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc cggcatgccg
     5881 ccctttcgtc ttcaagaatt aattcccaat tccccaggca tcaaataaaa cgaaaggctc
     5941 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
     6001 ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg
     6061 c
//
