LOCUS       sequence-297209-        3519 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression and purification of 4xTR-TUBE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3519)
  AUTHORS   Tsuchiya H, Burana D, Ohtake F, Arai N, Kaiho A, Komada M, Tanaka K,
            Saeki Y
  TITLE     Ub-ProT reveals global length and composition of protein 
            ubiquitylation in cells.
  JOURNAL   Nat Commun. 2018 Feb 6;9(1):524. doi: 10.1038/s41467-018-02869-x.
  PUBMED    29410401
REFERENCE   2  (bases 1 to 3519)
  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: "Nat 
            Commun."; date: "2018-02-6"; volume: "9(1):524. doi"; pages: " 
            10.1038/s41467-018-02869-x"
COMMENT     Sequence Label: pRSET-4xTR-TUBE
FEATURES             Location/Qualifiers
     source          1..3519
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             112..129
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             133..165
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     terminator      934..981
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1078..1533
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1165..1184)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1375..1396
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1559..1663
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1664..1732,1733..2524)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1882..1901)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2695..3283
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3184..3203
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3437..3454
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcggggttg tcatcatcat
      121 catcatcatg gtatggctag catgactggt ggacagcaaa tgggtgatat cggcggcggt
      181 ggttcgggag gtggggtaaa tccgcagctg cagaatcctg aggtcgcgtt tcaacagcag
      241 ttggagcaat tgagcgcaat ggggttctta aatgcggaag cgaaccttca ggccctcatt
      301 gccaccggtg gggatattaa tgcggccatt gaagccctgc tgggttcgca gccatccggc
      361 ggcggcgggt ctggtggagg ggtgaaccct cagttacaga atccggaagt cgcgtttcag
      421 caacaattag agcagttaag cgccatgggc tttcttaacg cggaagcgaa cttgcaggcc
      481 ctgatcgcga ctggtggaga tattaatgca gccatcgagg cacttctggg gagccagccc
      541 agtggcggcg gcgggagcgg tggaggtgtg aacccgcagt tgcaaaaccc ggaagtggcg
      601 ttccaacagc agctggaaca actgtcggcc atgggttttc tgaacgcaga agctaacctc
      661 caagcgctga ttgctaccgg tggcgatatt aacgctgcga tcgaagcact gctcggctca
      721 cagccaagtg gcggtggcgg cagcggtggc ggcgttaatc cgcaactgca aaaccccgaa
      781 gttgcgtttc agcagcaact ggagcagctg tccgcaatgg gcttcctgaa tgccgaagca
      841 aatctgcagg ctctcattgc tacgggtggc gacatcaacg ctgccatcga agcgctgctg
      901 ggctctcagc cgtcaggagg tggtggatcc taactagcat aaccccttgg ggcctctaaa
      961 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggatctgg cgtaatagcg
     1021 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     1081 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     1141 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     1201 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     1261 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     1321 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     1381 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     1441 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     1501 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     1561 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     1621 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     1681 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     1741 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     1801 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     1861 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     1921 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     1981 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     2041 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     2101 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     2161 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     2221 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     2281 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     2341 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     2401 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     2461 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     2521 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     2581 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     2641 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2701 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2761 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2821 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     2881 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2941 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3001 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3061 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3121 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3181 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3241 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3301 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3361 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3421 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     3481 accgcctctc cccgcgcgtt ggccgattca ttaatgcag
//
