LOCUS       sequence-222070-        4811 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  translational release factor 2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4811)
  AUTHORS   Villarreal F, Contreras-Llano LE, Chavez M, Ding Y, Fan J, Pan T, 
            Tan C
  TITLE     Synthetic microbial consortia enable rapid assembly of pure 
            translation machinery.
  JOURNAL   Nat Chem Biol. 2018 Jan;14(1):29-35. doi: 10.1038/nchembio.2514. 
            Epub 2017 Nov 13.
  PUBMED    29131146
REFERENCE   2  (bases 1 to 4811)
  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/nchembio.2514"; journalName: "Nat Chem Biol"; date: 
            "2018-01"; volume: "14"; issue: "1"; pages: "29-35"
COMMENT     Sequence Label: B3-4
FEATURES             Location/Qualifiers
     source          1..4811
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..111
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        92..110
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             150..172
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             182..199
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1371..1389)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1385..1432
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2041..2991)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      3039..3261
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3753..3847)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3878..4672)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3989..4008)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4599..4618
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 gtctaagaaa ccattattat catgacatta acctataaaa ataggcggat cacgaggccc
       61 tttcgtcttc acctcgatga tcccgcgaaa ttaatacgac tcactatagg ggaattgaga
      121 gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga gatataccat
      181 gcatcatcat caccatcatt tatttgaaat taatccggta aataatcgca ttcaggacct
      241 cacggaacgc tccgacgttc ttagggggta tcttgactac gacgccaaga aagagcgtct
      301 ggaagaagta aacgccgagc tggaacagcc ggatgtctgg aacgaacccg aacgcgcaca
      361 ggcgctgggt aaagagcgtt cctccctcga agccgttgtc gacaccctcg accaaatgaa
      421 acaggggctg gaagatgttt ctggtctgct ggaactggct gtagaagctg acgacgaaga
      481 aacctttaac gaagccgttg ctgaactcga cgccctggaa gaaaaactgg cgcagcttga
      541 gttccgccgt atgttctctg gcgaatatga cagcgccgac tgctacctcg atattcaggc
      601 ggggtctggc ggtacggaag cacaggactg ggcgagcatg cttgagcgta tgtatctgcg
      661 ctgggcagaa tcgcgtggtt tcaaaactga aatcatcgaa gagtcggaag gtgaagtggc
      721 gggtattaaa tccgtgacga tcaaaatctc cggcgattac gcttacggct ggctgcgtac
      781 agaaaccggc gttcaccgcc tggtgcgtaa aagcccgttt gactccggcg gtcgtcgcca
      841 cacgtcgttc agctccgcgt ttgtttatcc ggaagttgat gatgatattg atatcgaaat
      901 caacccggcg gatctgcgca ttgacgttta tcgcacgtcc ggcgcgggcg gtcagcacgt
      961 taaccgtacc gaatctgcgg tgcgtattac ccacatcccg accgggatcg tgacccagtg
     1021 ccagaacgac cgttcccagc acaagaacaa agatcaggcc atgaagcaga tgaaagcgaa
     1081 gctttatgaa ctggagatgc agaagaaaaa tgccgagaaa caggcgatgg aagataacaa
     1141 atccgacatc ggctggggca gccagattcg ttcttatgtc cttgatgact cccgcattaa
     1201 agatctgcgc accggggtag aaacccgcaa cacgcaggcc gtgctggacg gcagcctgga
     1261 tcaatttatc gaagcaagtt tgaaagcagg gttatattaa agcagaggcc atatgctcga
     1321 ggatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca
     1381 ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg
     1441 aggaactata tccggatatc ccgcaagagg cccggcagta ccggcataac caagcctatg
     1501 cctacagcat ccagggccta gggtacgggt tttgctgccc gcaaacgggc tgttctggtg
     1561 ttgctagttt gttatcagaa tcgcagatcc ggcttcagcc ggtttgccgg ctgaaagcgc
     1621 tatttcttcc agaattgcca tgattttttc cccacgggag gcgtcactgg ctcccgtgtt
     1681 gtcggcagct ttgattcgat aagcagcatc gcctgtttca ggctgtctat gtgtgactgt
     1741 tgagctgtaa caagttgtct caggtgttca atttcatgtt ctagttgctt tgttttactg
     1801 gtttcacctg ttctattagg tgttacatgc tgttcatctg ttacattgtc gatctgttca
     1861 tggtgaacag ctttgaatgc accaaaaact cgtaaaagct ctgatgtatc tatctttttt
     1921 acaccgtttt catctgtgca tatggacagt tttccctttg atatgtaacg gtgaacagtt
     1981 gttctacttt tgtttgttag tcttgatgct tcactgatag atacaagagc cataagaacc
     2041 tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag
     2101 ccatgagaac gaaccattga gatcatactt actttgcatg tcactcaaaa attttgcctc
     2161 aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt
     2221 atgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg
     2281 gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa
     2341 cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa
     2401 atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt
     2461 ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt
     2521 tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc
     2581 aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg
     2641 caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt
     2701 ccactggaaa atctcaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat
     2761 cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat
     2821 ctgagcgtat tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat
     2881 cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc
     2941 atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa
     3001 ttggtctagg tgattttaat cactatacca attgagatgg gctagtcaat gataattact
     3061 agtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact
     3121 tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta
     3181 tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga
     3241 tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc
     3301 gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac
     3361 cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc tccgaccatc aggcacctga
     3421 gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac ggctctggca gtgaatgggg
     3481 gtaaatggca ctacaggcgc cttttatgga ttcatgcaag gaaactaccc ataatacaag
     3541 aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg tctgctatgt ggtgctatct
     3601 gactttttgc tgttcagcag ttcctgccct ctgattttcc agtctgacca cttcggatta
     3661 tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggt atcatcaaca
     3721 ggcttacccg tcttactgtc cctagtgctt ggattctcac caataaaaaa cgcccggcgg
     3781 caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg atctatcaac
     3841 aggagtccaa gcgagctctc gaaccccaga gtcccgctca gaagaactcg tcaagaaggc
     3901 gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg aggaagcggt
     3961 cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct atgtcctgat
     4021 agcggtccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg ccattttcca
     4081 ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg ccgtcgggca
     4141 tgcgcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc tcttcgtcca
     4201 gatcatcctg atcgacaaga ccggcttcca tccgagtacg tgctcgctcg atgcgatgtt
     4261 tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt atgcagccgc cgcattgcat
     4321 cagccatgat ggatactttc tcggcaggag caaggtgaga tgacaggaga tcctgccccg
     4381 gcacttcgcc caatagcagc cagtcccttc ccgcttcagt gacaacgtcg agcacagctg
     4441 cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc tgcctcgtcc tgcagttcat
     4501 tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc gctgacagcc
     4561 ggaacacggc ggcatcagag cagccgattg tctgttgtgc ccagtcatag ccgaatagcc
     4621 tctccaccca agcggccgga gaacctgcgt gcaatccatc ttgttcaatc atgcgaaacg
     4681 atcctcatcc tgtctcttga tcagatcttg atcccctgcg ccatcagatc cttggcggca
     4741 agaaagccat ccagtttact ttgcagggct tcccaacctt accagagggc gccccagctg
     4801 gcaattccga c
//
