LOCUS       sequence-357023-        5323 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of N-terminally 6His tagged A1-LCD with seven 
            Arg residues added, twelve Glu residues added (LCD: low-complexity 
            domain, aa 186-320).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5323)
  AUTHORS   Bremer A, Farag M, Borcherds WM, Peran I, Martin EW, Pappu RV, 
            Mittag T
  TITLE     Deciphering how naturally occurring sequence features impact the 
            phase behaviours of disordered prion-like domains.
  JOURNAL   Nat Chem. 2022 Feb;14(2):196-207. doi: 10.1038/s41557-021-00840-w. 
            Epub 2021 Dec 20.
  PUBMED    34931046
REFERENCE   2  (bases 1 to 5323)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41557-021-00840-w"; journalName: "Nat Chem"; date: 
            "2022-02"; volume: "14"; issue: "2"; pages: "196-207"
COMMENT     Sequence Label: A1-LCD+7R+12D
FEATURES             Location/Qualifiers
     source          1..5323
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..40
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             71..93
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             113..130
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             131..154
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     protein_bind    332..356
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             359..379
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     protein_bind    complement(792..816)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(869..887)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      883..930
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1296..1301,1302..1318,1319..1324))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1351..1369)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1437..1541
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1542..1610,1611..2402)
                     /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(1760..1779)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2573..3161
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3062..3081
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3315..3332
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3347..3486
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3412..3431)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3572..3594
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3588..3779)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5241..5260
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 agatctcgat cccgcgaaat taatacgact cactataggg agaccacaac ggtttccctc
       61 tagaaataat tttgtttaac tttaagaagg agatatacat atgtcgtact accatcacca
      121 tcaccatcac ctggaagtgc tgtttcaggg cccggatacc tataaactgg tgattgtgct
      181 gaacggcacc acctttacct ataccaccga agcggtggat gcggcgaccg cggaaaaagt
      241 gtttaaacag tatgcgaacg atgcgggcgt ggatggcgaa tggacctatg atgcggcgac
      301 caaaaccttt accgtgaccg aactcgaatc aacaagtttg tacaaaaaag caggcttcga
      361 gaacctgtat tttcagggta gcatggcaag cgcagatagc agccagcgtg atcgtgatga
      421 tcgtggtaat tttggtgatg gtcgcggtgg tggttttggt ggtaatgata attttggccg
      481 tggtggcaat tttagcgatc gcggtggctt tggcggtagc cgtggtgatg gccgttatgg
      541 cggtgatggt gatcgttata atggttttgg caatgacggt cgcaactttg gtggtggtgg
      601 cagctataac gattttggca actataataa ccagagcagc aacttcgatc cgatgaaagg
      661 cggtaacttt cgtgatcgta gcagcggtcc gtatgaccgt ggtggtcagt attttgccaa
      721 accgcgtaat caaggtggct atggtggtag tagcagcagc cgtagctatg gtagcgatcg
      781 tcgtttttaa gacccagctt tcttgtacaa agtggttgat tcgaggctgc taacaaagcc
      841 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
      901 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatatcca
      961 caggacgggt gtggtcgcca tgatcgcgta gtcgatagtg gctccaagta gcgaagcgag
     1021 caggactggg cggcggccaa agcggtcgga cagtgctccg agaacgggtg cgcatagaaa
     1081 ttgcatcaac gcatatagcg ctagcagcac gccatagtga ctggcgatgc tgtcggaatg
     1141 gacgatatcc cgcaagaggc ccggcagtac cggcataacc aagcctatgc ctacagcatc
     1201 cagggtgacg gtgccgagga tgacgatgag cgcattgtta gatttcatac acggtgcctg
     1261 actgcgttag caatttaact gtgataaact accgcattaa agcttatcga tgataagctg
     1321 tcaaacatga gaattcttga agacgaaagg gcctcgtgat acgcctattt ttataggtta
     1381 atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg
     1441 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     1501 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     1561 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     1621 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     1681 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     1741 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag
     1801 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     1861 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     1921 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     1981 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     2041 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa
     2101 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     2161 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     2221 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     2281 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     2341 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     2401 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     2461 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     2521 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     2581 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     2641 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     2701 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     2761 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     2821 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     2881 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     2941 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     3001 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     3061 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     3121 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     3181 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     3241 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     3301 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     3361 ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg
     3421 ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg
     3481 gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     3541 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     3601 ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc
     3661 gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt
     3721 aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact
     3781 gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag
     3841 aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag
     3901 ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc
     3961 cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg
     4021 cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca
     4081 cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg
     4141 cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag
     4201 cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggcca ggacccaacg
     4261 ctgcccgaga tgcgccgcgt gcggctgctg gagatggcgg acgcgatgga tatgttctgc
     4321 caagggttgg tttgcgcatt cacagttctc cgcaagaatt gattggctcc aattcttgga
     4381 gtggtgaatc cgttagcgag gtgccgccgg cttccattca ggtcgaggtg gcccggctcc
     4441 atgcaccgcg acgcaacgcg gggaggcaga caaggtatag ggcggcgcct acaatccatg
     4501 ccaacccgtt ccatgtgctc gccgaggcgg cataaatcgc cgtgacgatc agcggtccaa
     4561 tgatcgaagt taggctggta agagccgcga gcgatccttg aagctgtccc tgatggtcgt
     4621 catctacctg cctggacagc atggcctgca acgcgggcat cccgatgccg ccggaagcga
     4681 gaagaatcat aatggggaag gccatccagc ctcgcgtcgc gaacgccagc aagacgtagc
     4741 ccagcgcgtc ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg
     4801 cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca
     4861 ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg
     4921 ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag
     4981 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
     5041 gatcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg
     5101 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
     5161 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
     5221 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
     5281 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcg
//
