LOCUS       sequence-376751-        5081 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses His8-EWS_RRM-RGG(359-513) with a TEV site for removal of 
            the His8 tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5081)
  AUTHORS   Selig EE, Bhura R, White MR, Akula S, Hoffman RD, Tovar CN, Xu X, 
            Booth RE, Libich DS
  TITLE     Biochemical and biophysical characterization of the nucleic acid 
            binding properties of the RNA/DNA binding protein EWS.
  JOURNAL   Biopolymers. 2023 May;114(5):e23536. doi: 10.1002/bip.23536. Epub 
            2023 Mar 17.
  PUBMED    36929870
REFERENCE   2  (bases 1 to 5081)
  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: 
            "Biopolymers."; date: "2023-05"; volume: "114(5):e23536. doi"; 
            pages: " 10.1002/bip.23536. Epub 2023 Mar 17"
COMMENT     Sequence Label: pAG8Ha-EWS_RRM-RGG(359-513)
FEATURES             Location/Qualifiers
     source          1..5081
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..15,16..33,34..39)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(214..261)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     257..275
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(797..817)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(824..847)
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     RBS             862..884
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(915..934)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(916..934)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2636..2827
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2821..2843)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2929..3068
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2984..3003
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3083..3100)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3254..3842)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3334..3353)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4013..4804,4805..4873))
                     /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     4636..4655
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4874..4978)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5046..5064
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ttctcatgtt tgacagctta tcatccgata agctttaatg cggtagttta tcacagttaa
       61 attgctaacg cagtcaggca ccgtgtatga aatctaacaa tgcgctcatc gtcatcctcg
      121 gcaccgtcac cctggatgct gtaggcatag gcttggttat gccggtactg ccgggcctct
      181 tgcgggatat ccggatatag ttcctccttt cagcaaaaaa cccctcaaga cccgtttaga
      241 ggccccaagg ggttatgcta gttattgctc agcggtggca gcagccaact cagcttcctt
      301 tcgggctttg ttagcagccg gatcctcagc caccgccgct cgggttacca cggctgccac
      361 gcggaccacg cggcggaaaa ccgccacggt caccgccacg accgcccata cgacccatcg
      421 gaccgcccgg accgcccgga ccgcccggac cgccacgcag tggcggcggc atgccacgac
      481 cctcacgcgg cggcagaccg ccacgcatgc tgttcatcgg cggtttctta cgcgccaggc
      541 tcactttcag cttgctaccc tgaaaatctt tgccgtcgaa ccattcaacc gccgccttcg
      601 cggtcggcgg gtcctcgtag ctcacggtcg cgtcaccttt cggcttgccg gtttctttat
      661 ccaggtaaat gtggatcatc ggttgaccgg tacgtttgtt catcttaacc acgccgcact
      721 gcttaaagaa atccgccaga tcgtccaggg taacgctgtc gttcagaccc tgcacgtaga
      781 tcgcgctgtt atcggcgccc tgaaaataaa gattctcgct agcatggtga tggtggtggt
      841 gatggtggtc catatggtat atctccttct taaagttaaa caaaattatt tctagaggga
      901 aaccgttgtg gtctccctat agtgagtcgt attaatttcg cgggatcgag atctcgatcc
      961 tctacgccgg acgcatcgtg gccggcatca ccggcgctac aggtgcggtt gcttgcgcct
     1021 atatcgccga cccggcatca ccggcgctac aggtgcggtt gcttgcgcct atatcgccga
     1081 catcaccgat ggggaagatc gggctcgcca cttcgggctc atgagcgctt gtttcggcgt
     1141 gggtatggtg gcaggccccg tggccgtggg actgttgggc tccatctcct tgcatgggga
     1201 ctgttgggct ccatctcctt gcatgggggg actgttgggc tccatctcct tgcatgcacc
     1261 attccttgcg gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca
     1321 ggagtcgcat aagggagagc gtcaaccgat gcccttgaga gccttcaacc cagtcagctc
     1381 cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat
     1441 gcaactcgta ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg accgctttcg
     1501 ctggagcgcg acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc
     1561 tcaagccttc gtcactggtc ccgccaccaa acgtttcggc gagaagcagg ccattatcgc
     1621 cggcatggcg gccgacgcgc tgggctacgt cttgctggcg ttcgcgacgc gaggctggat
     1681 ggccttcccc attatgattc ttctcgcttc cggcggcatc gggatgcccg cgttgcaggc
     1741 catgctgtcc aggcaggtag atgacgacca tcagggacag cttcaaggat cgctcgcggc
     1801 tcttaccagc ctaacttcga tcattggacc gctgatcgtc acggcgattt atgccgcctc
     1861 ggcgagcaca tggaacgggt tggcatggat tgtaggtgcc gccctatacc ttgtcatgga
     1921 ttgtaggtgc cgccctatac cttgtctgcc tccccgcgtt gcgtcgcggt gcatggagcc
     1981 gggccacctc gacctgaatg gaagccggcg gcacctcgct aacggattca ccactccaag
     2041 aattggagcc aatcaattct tgcggagaac tgtgaatgcg caaaccaacc cttggcagaa
     2101 catatccatc gcgtccgcca tctccagcag ccgcacgcgg cgcatctcgg gcagcgttgg
     2161 gtcctggcca cgggtgtgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg
     2221 gggttgcctt actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg
     2281 ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc
     2341 gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc
     2401 aggatgctgc tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg
     2461 accctgagtg atttttctct ggtcccgccg catccatacc gccagttgtt taccctcacg
     2521 acgttccagt aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg
     2581 tttcatcggt atcattaccc ccatgaacag aaatccccct tacacggagg catcagtgac
     2641 caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga cattaacgct
     2701 tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg aatcgcttca
     2761 cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg acggtgaaaa
     2821 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     2881 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     2941 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     3001 gtactgagag tgcaccattg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     3061 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3121 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3181 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3241 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3301 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3361 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3421 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     3481 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     3541 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     3601 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     3661 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     3721 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3781 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3841 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3901 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3961 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4021 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4081 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4141 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4201 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4261 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4321 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4381 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4441 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     4501 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     4561 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     4621 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     4681 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     4741 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     4801 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     4861 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     4921 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     4981 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     5041 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga a
//
