LOCUS       sequence-322701-        7271 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of N. furzeri DDX5 in E. coli .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7271)
  AUTHORS   Harel I, Chen YR, Ziv I, Singh PP, Heinzer D, Navarro Negredo P, 
            Goshtchevsky U, Wang W, Astre G, Moses E, McKay A, Machado BE, 
            Hebestreit K, Yin S, Sanchez Alvarado A, Jarosz DF, Brunet A
  TITLE     Identification of protein aggregates in the aging vertebrate brain 
            with prion-like and phase-separation properties.
  JOURNAL   Cell Rep. 2024 Jun 25;43(6):112787. doi: 
            10.1016/j.celrep.2023.112787. Epub 2024 May 28.
  PUBMED    38810650
REFERENCE   2  (bases 1 to 7271)
  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"; journalName: "Cell Rep.";
            date: "2024-06-25"; volume: "43(6):112787. doi"; pages: " 
            10.1016/j.celrep.2023.112787. Epub 2024 May 28"
COMMENT     Sequence Label: pDEST17-His10-DDX5(kf)-SNAP
FEATURES             Location/Qualifiers
     source          1..7271
                     /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..142
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             155..166
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    188..212
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2133..2157)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2159..2182
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2201..2746
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="optimized for bacterial expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHEIKLLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYQQLAALAGNPAATAAVKTALSGNPVPILIPCHRVVSSSGAVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     primer_bind     complement(2814..2832)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2828..2875
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(3241..3246,3247..3263,3264..3269))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(3296..3314)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3382..3486
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3487..3555,3556..4347)
                     /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(3705..3724)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4518..5106
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5007..5026
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5260..5277
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5292..5434
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5360..5379)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5520..5542
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5536..5727)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     7189..7208
                     /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 catcatcatc acagcagcgg ccatatcgaa ggtcgtcata tggctagcct
      181 cgaatcaaca agtttgtaca aaaaagcagg cttcgaagga gataacaaaa tgcctggatt
      241 tgcagacaga gaccgcggca gagatagagg gtatgggggt ggcccccctc ggtttggtgg
      301 cggtggggga ggaggcggca acagaggagg acctctgctg ggaaagtttg gaaaccctgg
      361 tgagaggctg cgcaagaagc actggaacct tgatgaacta ccaaaatttc aaaagaattt
      421 ctaccaggaa cacccagatg tcacacacag accactgcag gaggttgacc agtaccgcag
      481 aaacaaagaa gttacagtta aaggcagaga ttgtcccaaa ccgattgtaa aattccatga
      541 agctgctttt cccaactacg tcatggatgt cattgtcaaa cagaactgga ctgaaccaac
      601 tccgatccag tctcaggggt ggcccattgt cctgagtgga aaagacatgg tcggcatcgc
      661 tcaaacgggg tctgggaaga ctttggcata tcttctgcct gcagtcgttc acatccaaca
      721 tcagcccttc ttggagcacg gagacgggcc tatttgcttg gtgctggcac caacccgtga
      781 gctggctcag caggtgcaac aggtggctgc tgagtacggc agggcctccc gtctcaagag
      841 tacatgcatc tatggtggtg ctcccaaagg accacaaatc cgggaccttg agaggggtgt
      901 tgagatttgc atcgccaccc caggtcgcct cattgatttc ctggaatgtg gtaaaactaa
      961 tttgcgtcgt tgcacctatc tagtgctgga tgaagctgac cgcatgctgg acatgggctt
     1021 tgaacctcag atccgcaaga tagtggatca aatacggcca gaccgtcaga ctctgatgtg
     1081 gagtgccacc tggcctaaag aagttcgtca gctggctgag gacttcttga aggattacgt
     1141 ccagatcaac attggcgccc tgcagctcag cgccaatcat aacatcctgc aaatagtaga
     1201 tgtttgcaat gacttagaaa aggaagacaa actgattcgg ttgctggagg agatcatgag
     1261 tgaaaaggaa aacaagacca tcatttttgt ggagaccaaa aggcgttgtg atgagctcac
     1321 caggaggatg agaagagacg gatggcctgc aatgggaatc catggagata agagccagca
     1381 ggagagggac tgggtcctta acgagttcag atttggcaaa gctccgattc tcattgctac
     1441 agacgtggcc tcccggggct tagatgtgga ggatgtgaaa tttgtcatca attatgacta
     1501 ccctaactcc tctgaggatt atatccaccg cattggacgc acagcccgaa gtcaaaaaac
     1561 gggcacggcc tacaccttct tcacccccaa caacatgaaa caggcaggtg acctgatctc
     1621 tgtgctccgt gaagccaacc aggccatcaa ccccaaactg atccagatgg ctgaggacag
     1681 aggaggtcgc ggtagggggg gaagaggtgg ctacaaggat gaccgtcgcg ataggtattc
     1741 tgggggtggg aggagtaatt ttggcggtag tggctttaga gacagggacc atgacagagg
     1801 tttcggcaat gggccgaaga ccacatttgg tggcagcaag gcccaaaata gtggcaatta
     1861 tggtggcaac agtggtaact cttgtggtag ctatggcaac aataactaca acaacagtaa
     1921 tggacagggt agttttagtg gtccacctaa ccaaggtggc tttgggaacc agggcttcca
     1981 agcccctcct caatttggtg gcatgcagag ggctgcacag aatggtgtta atcatccacc
     2041 atttcctttt aacgctcagc cacccccgcc acagaaccag cagccccctc caccaatggt
     2101 gccctaccca attccaccac cgtttccaca agacccagct ttcttgtaca aagtggttga
     2161 ctacaaggat gacgatgaca agcttaccgg tgagttcgct atggacaaag attgcgaaat
     2221 gaaacgtacc accctggata gcccgctggg caaactggaa ctgagcggct gcgaacaggg
     2281 cctgcatgaa attaaactgc tgggtaaagg caccagcgcg gccgatgcgg ttgaagttcc
     2341 ggccccggcc gccgtgctgg gtggtccgga accgctgatg caggcgaccg cgtggctgaa
     2401 cgcgtatttt catcagccgg aagcgattga agaatttccg gttccggcgc tgcatcatcc
     2461 ggtgtttcag caggagagct ttacccgtca ggtgctgtgg aaactgctga aagtggttaa
     2521 atttggcgaa gtgattagct atcagcagct ggcggccctg gcgggtaatc cggcggccac
     2581 cgccgccgtt aaaaccgcgc tgagcggtaa cccggtgccg attctgattc cgtgccatcg
     2641 tgtggttagc tctagcggtg cggttggcgg ttatgaaggt ggtctggcgg tgaaagagtg
     2701 gctgctggcc catgaaggtc atcgtctggg taaaccgggt ctgggacctg caggctgata
     2761 aggattcgag gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
     2821 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     2881 aggaggaact atatccggat atccacagga cgggtgtggt cgccatgatc gcgtagtcga
     2941 tagtggctcc aagtagcgaa gcgagcagga ctgggcggcg gccaaagcgg tcggacagtg
     3001 ctccgagaac gggtgcgcat agaaattgca tcaacgcata tagcgctagc agcacgccat
     3061 agtgactggc gatgctgtcg gaatggacga tatcccgcaa gaggcccggc agtaccggca
     3121 taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat
     3181 tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc
     3241 attaaagctt atcgatgata agctgtcaaa catgagaatt cttgaagacg aaagggcctc
     3301 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     3361 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     3421 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     3481 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     3541 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     3601 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     3661 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     3721 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     3781 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     3841 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     3901 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     3961 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     4021 acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     4081 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     4141 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     4201 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     4261 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     4321 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     4381 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     4441 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     4501 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     4561 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     4621 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     4681 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     4741 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     4801 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     4861 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     4921 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     4981 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     5041 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     5101 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     5161 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     5221 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     5281 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     5341 atatatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     5401 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     5461 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     5521 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     5581 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     5641 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     5701 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     5761 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     5821 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     5881 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     5941 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     6001 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     6061 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     6121 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     6181 catgcgcacc cgtggccagg acccaacgct gcccgagatg cgccgcgtgc ggctgctgga
     6241 gatggcggac gcgatggata tgttctgcca agggttggtt tgcgcattca cagttctccg
     6301 caagaattga ttggctccaa ttcttggagt ggtgaatccg ttagcgaggt gccgccggct
     6361 tccattcagg tcgaggtggc ccggctccat gcaccgcgac gcaacgcggg gaggcagaca
     6421 aggtataggg cggcgcctac aatccatgcc aacccgttcc atgtgctcgc cgaggcggca
     6481 taaatcgccg tgacgatcag cggtccagtg atcgaagtta ggctggtaag agccgcgagc
     6541 gatccttgaa gctgtccctg atggtcgtca tctacctgcc tggacagcat ggcctgcaac
     6601 gcgggcatcc cgatgccgcc ggaagcgaga agaatcataa tggggaaggc catccagcct
     6661 cgcgtcgcga acgccagcaa gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg
     6721 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     6781 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     6841 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     6901 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     6961 gggttgaagg ctctcaaggg catcggtcga tcgacgctct cccttatgcg actcctgcat
     7021 taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc
     7081 atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
     7141 gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc
     7201 gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc
     7261 gtagaggatc g
//
