LOCUS       sequence-222063-        4318 bp    DNA     circular SYN 14-APR-2025
DEFINITION  ribosome recycling factor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4318)
  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 4318)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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: B4-5
FEATURES             Location/Qualifiers
     source          1..4318
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     40..58
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     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             737..754
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(878..896)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      892..939
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1548..2498)
                     /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      2546..2768
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3260..3354)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3385..4179)
                     /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(3496..3515)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4106..4125
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc
       61 tttcgtcttc acctcgatga tcccgcgaaa ttaatacgac tcactatagg ggaattgaga
      121 gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga gatataccat
      181 gcatattagc gatatcagaa aagatgctga agtacgcatg gacaaatgcg tagaagcgtt
      241 caaaacccaa atcagcaaaa tacgcacggg tcgtgcttct cccagcctgc tggatggcat
      301 tgtcgtggaa tattacggca cgccgacgcc gctgcgtcag ctggcaagcg taacggtaga
      361 agattcccgt acactgaaaa tcaacgtgtt tgatcgttca atgtctccgg ccgttgaaaa
      421 agcgattatg gcgtccgatc ttggcctgaa cccgaactct gcgggtagcg acatccgtgt
      481 tccgctgccg ccgctgacgg aagaacgtcg taaagatctg accaaaatcg ttcgtggtga
      541 agcagaacaa gcgcgtgttg cagtacgtaa cgtgcgtcgt gacgcgaacg acaaagtgaa
      601 agcactgttg aaagataaag agatcagcga agacgacgat cgccgttctc aggacgatgt
      661 acagaaactg actgatgctg caatcaagaa aattgaagcg gcgctggcag acaaagaagc
      721 agaactgatg cagttccatc atcatcacca tcattaaagc cttctatcgt tgccaaactg
      781 gcggcgttgt ccgagcagga atattaaagc agaggccata tgctcgagga tccggctgct
      841 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
      901 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc
      961 ggatatcccg caagaggccc ggcagtaccg gcataaccaa gcctatgcct acagcatcca
     1021 gggcctaggg tacgggtttt gctgcccgca aacgggctgt tctggtgttg ctagtttgtt
     1081 atcagaatcg cagatccggc ttcagccggt ttgccggctg aaagcgctat ttcttccaga
     1141 attgccatga ttttttcccc acgggaggcg tcactggctc ccgtgttgtc ggcagctttg
     1201 attcgataag cagcatcgcc tgtttcaggc tgtctatgtg tgactgttga gctgtaacaa
     1261 gttgtctcag gtgttcaatt tcatgttcta gttgctttgt tttactggtt tcacctgttc
     1321 tattaggtgt tacatgctgt tcatctgtta cattgtcgat ctgttcatgg tgaacagctt
     1381 tgaatgcacc aaaaactcgt aaaagctctg atgtatctat cttttttaca ccgttttcat
     1441 ctgtgcatat ggacagtttt ccctttgata tgtaacggtg aacagttgtt ctacttttgt
     1501 ttgttagtct tgatgcttca ctgatagata caagagccat aagaacctca gatccttccg
     1561 tatttagcca gtatgttctc tagtgtggtt cgttgttttt gcgtgagcca tgagaacgaa
     1621 ccattgagat catacttact ttgcatgtca ctcaaaaatt ttgcctcaaa actggtgagc
     1681 tgaatttttg cagttaaagc atcgtgtagt gtttttctta gtccgttatg taggtaggaa
     1741 tctgatgtaa tggttgttgg tattttgtca ccattcattt ttatctggtt gttctcaagt
     1801 tcggttacga gatccatttg tctatctagt tcaacttgga aaatcaacgt atcagtcggg
     1861 cggcctcgct tatcaaccac caatttcata ttgctgtaag tgtttaaatc tttacttatt
     1921 ggtttcaaaa cccattggtt aagcctttta aactcatggt agttattttc aagcattaac
     1981 atgaacttaa attcatcaag gctaatctct atatttgcct tgtgagtttt cttttgtgtt
     2041 agttctttta ataaccactc ataaatcctc atagagtatt tgttttcaaa agacttaaca
     2101 tgttccagat tatattttat gaattttttt aactggaaaa gataaggcaa tatctcttca
     2161 ctaaaaacta attctaattt ttcgcttgag aacttggcat agtttgtcca ctggaaaatc
     2221 tcaaagcctt taaccaaagg attcctgatt tccacagttc tcgtcatcag ctctctggtt
     2281 gctttagcta atacaccata agcattttcc ctactgatgt tcatcatctg agcgtattgg
     2341 ttataagtga acgataccgt ccgttctttc cttgtagggt tttcaatcgt ggggttgagt
     2401 agtgccacac agcataaaat tagcttggtt tcatgctccg ttaagtcata gcgactaatc
     2461 gctagttcat ttgctttgaa aacaactaat tcagacatac atctcaattg gtctaggtga
     2521 ttttaatcac tataccaatt gagatgggct agtcaatgat aattactagt ccttttcctt
     2581 tgagttgtgg gtatctgtaa attctgctag acctttgctg gaaaacttgt aaattctgct
     2641 agaccctctg taaattccgc tagacctttg tgtgtttttt ttgtttatat tcaagtggtt
     2701 ataatttata gaataaagaa agaataaaaa aagataaaaa gaatagatcc cagccctgtg
     2761 tataactcac tactttagtc agttccgcag tattacaaaa ggatgtcgca aacgctgttt
     2821 gctcctctac aaaacagacc ttaaaaccct aaaggcttaa gtagcaccct cgcaagctcg
     2881 ggcaaatcgc tgaatattcc ttttgtctcc gaccatcagg cacctgagtc gctgtctttt
     2941 tcgtgacatt cagttcgctg cgctcacggc tctggcagtg aatgggggta aatggcacta
     3001 caggcgcctt ttatggattc atgcaaggaa actacccata atacaagaaa agcccgtcac
     3061 gggcttctca gggcgtttta tggcgggtct gctatgtggt gctatctgac tttttgctgt
     3121 tcagcagttc ctgccctctg attttccagt ctgaccactt cggattatcc cgtgacaggt
     3181 cattcagact ggctaatgca cccagtaagg cagcggtatc atcaacaggc ttacccgtct
     3241 tactgtccct agtgcttgga ttctcaccaa taaaaaacgc ccggcggcaa ccgagcgttc
     3301 tgaacaaatc cagatggagt tctgaggtca ttactggatc tatcaacagg agtccaagcg
     3361 agctctcgaa ccccagagtc ccgctcagaa gaactcgtca agaaggcgat agaaggcgat
     3421 gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc
     3481 gccaagctct tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac
     3541 acccagccgg ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg
     3601 caagcaggca tcgccatggg tcacgacgag atcctcgccg tcgggcatgc gcgccttgag
     3661 cctggcgaac agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc
     3721 gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc
     3781 gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga
     3841 tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa
     3901 tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc
     3961 cgtcgtggcc agccacgata gccgcgctgc ctcgtcctgc agttcattca gggcaccgga
     4021 caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc
     4081 atcagagcag ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc
     4141 ggccggagaa cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt
     4201 ctcttgatca gatcttgatc ccctgcgcca tcagatcctt ggcggcaaga aagccatcca
     4261 gtttactttg cagggcttcc caaccttacc agagggcgcc ccagctggca attccgac
//
