LOCUS       sequence-359564-        5096 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses the construct His8-GB1-EWS_171-264 with a TEV cleavage 
            site between GB1 and EWS_171-264.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5096)
  AUTHORS   Johnson CN, Xu X, Holloway SP, Libich DS
  TITLE     The (1)H, (15)N and (13)C resonance assignments of the 
            low-complexity domain from the oncogenic fusion protein EWS-FLI1.
  JOURNAL   Biomol NMR Assign. 2022 Apr;16(1):67-73. doi: 
            10.1007/s12104-021-10061-4. Epub 2022 Jan 7.
  PUBMED    34994941
REFERENCE   2  (bases 1 to 5096)
  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: 
            "10.1007/s12104-021-10061-4"; journalName: "Biomol NMR Assign"; 
            date: "2022-04"; volume: "16"; issue: "1"; pages: "67-73"
COMMENT     Sequence Label: pAG8H-GB1-EWS_171-264
FEATURES             Location/Qualifiers
     source          1..5096
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     51..70
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        51..69
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             101..123
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             138..161
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             363..383
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(725..743)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      739..786
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(961..966,967..984,985..990))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1017..1035)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1103..1207
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1208..1276,1277..2068)
                     /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(1426..1445)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2239..2827
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2728..2747
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2981..2998
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3013..3152
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3078..3097)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3238..3260
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3254..3445)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat taatacgact
       61 cactataggg agaccacaac ggtttccctc tagaaataat tttgtttaac tttaagaagg
      121 agatatacca tatggaccac catcaccacc accatcacca tgctagcatg cagtacaagc
      181 tggcgctgaa cggcaagacc ctgaaaggtg aaaccaccac cgaagcggtg gacgcggcga
      241 ccgcggagaa ggttttcaaa caatacgcga acgacaacgg tgtggatggc gaatggacct
      301 atgacgatgc gaccaaaacc ttcaccgtta ccgagggtag cgactacgat atttcgacca
      361 ccgaaaacct gtattttcag ggcacctatc cgcaggttcc gggcagctat ccgatgcagc
      421 cggttaccgc accgccgagc tatccgccga ccagctatag cagcacccag ccgaccagct
      481 atgatcagag cagctatagc cagcagaaca cctatggtca gccgagcagc tatggccagc
      541 agagcagcta tggtcaacaa agcagctatg gtcagcagcc gccgaccagc tatccgccgc
      601 agaccggtag ctatagccag gctccgagcc agtatagcca gcagagctct tcttatggcc
      661 agcagtgagg tctcggatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct
      721 gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt
      781 tttttgctga aaggaggaac tatatccgga tatcccgcaa gaggcccggc agtaccggca
      841 taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat
      901 tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc
      961 attaaagctt atcggatgat aagctgtcaa acatgagaat tcttgaagac gaaagggcct
     1021 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     1081 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     1141 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     1201 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     1261 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     1321 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     1381 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     1441 attatcccgt gttgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     1501 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     1561 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     1621 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     1681 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     1741 cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     1801 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     1861 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     1921 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     1981 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     2041 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     2101 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     2161 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     2221 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2281 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2341 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     2401 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2461 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2521 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2581 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2641 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2701 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2761 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     2821 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     2881 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     2941 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     3001 agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3061 caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc
     3121 cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg
     3181 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     3241 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt
     3301 aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca
     3361 gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa
     3421 gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg
     3481 gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac
     3541 atgcccggtt actggaacgt cgtgagggta aacaactggc ggtatggatg cggcgggacc
     3601 agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac
     3661 agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact
     3721 tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg
     3781 tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct
     3841 gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca
     3901 tgcacacccg tggccaggac ccaacgctgc ccgagatgcg ccgcgtgcgg ctgctggaga
     3961 tggcggacgc gatggatatg ttctgccaag ggttggtttg cgcattcaca gttctccgca
     4021 agaattgatt ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc cgccggcttc
     4081 cattcaggtc gaggtggccc ggctccatgc accgcgacgc aacgcgggga ggcagacaag
     4141 gtatagggcg gcacctacaa tccatgacaa ggtatagggc ggcacctaca atccatgcca
     4201 acccgttcca tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga
     4261 tcgaagttag gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat
     4321 ctacctgcct ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa
     4381 gaatcataat ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca
     4441 gcgcgtcggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     4501 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     4561 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     4621 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     4681 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggttgac
     4741 gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga
     4801 gcaccgccgc cgcaaggaat ggtgcatgca aggagatgga gcccaacagt ccccccatgc
     4861 aaggagatgg agcccaacag tccccatgca aggagatgga gcccaacagt cccacggcca
     4921 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
     4981 gatcttcccc atcggtgatg tcggcgatat aggcgcaagc aaccgcacct gtagcgccgg
     5041 tgatgccggg tcggcgatat aggcgcaagc aaccgcacct gtagcgccgg tgatgc
//
