LOCUS       sequence-217459-        6218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses K55M RPL28 from HIS3 yeast plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6218)
  AUTHORS   McGeachy AM, Meacham ZA, Ingolia NT
  TITLE     An Accessible Continuous-Culture Turbidostat for Pooled Analysis of 
            Complex Libraries.
  JOURNAL   ACS Synth Biol. 2019 Apr 19;8(4):844-856. doi: 
            10.1021/acssynbio.8b00529. Epub 2019 Apr 2.
  PUBMED    30908907
REFERENCE   2  (bases 1 to 6218)
  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: 
            "10.1021/acssynbio.8b00529"; journalName: "ACS Synth Biol"; date: 
            "2019-04-19- 19"; volume: "8"; issue: "4"; pages: "844-856"
COMMENT     Sequence Label: pRPL28(K55M) SpHIS5 (pNTI666)
FEATURES             Location/Qualifiers
     source          1..6218
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..61)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(215..803)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(295..314)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(974..1765,1766..1834))
                     /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     1597..1616
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1835..1939)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2007..2025
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2063..2085)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2185..2204
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2285..2303
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2285..2302
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     misc_feature    2349..2852
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     terminator      4590..4777
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(4645..4669)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            4852..6052
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        4852..5195
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5196..5849
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHTNQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMSNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     terminator      5855..6052
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(6156..6175)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6157..6175)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
       61 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
      121 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
      181 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
      241 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
      301 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
      361 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct
      421 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
      481 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
      541 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
      601 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
      661 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
      721 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
      781 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
      841 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
      901 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
      961 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     1021 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     1081 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     1141 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     1201 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     1261 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     1321 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     1381 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     1441 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     1501 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     1561 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     1621 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     1681 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     1741 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     1801 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     1861 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     1921 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     1981 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc
     2041 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     2101 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     2161 ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     2221 tggacatatt gtcgttagaa cgcggctaca attaatacat aaccttatgt atcatacaca
     2281 tacgatttag gtgacactat agaacgcggc cgccagctga agcttcgtac gctgcagggg
     2341 tccttttcat cacgtgctat aaaaataatt ataatttaaa ttttttaata taaatatata
     2401 aattaaaaat agaaagtaaa aaaagaaatt aaagaaaaaa tagtttttgt tttccgaaga
     2461 tgtaaaagac tctaggggga tcgccaacaa atactacctt ttatcttgct cttcctgctc
     2521 tcaggtatta atgccgaatt gtttcatctt gtctgtgtag aagaccacac acgaaaatcc
     2581 tgtgatttta cattttactt atcgttaatc gaatgtatat ctatttaatc tgcttttctt
     2641 gtctaataaa tatatatgta aagtacgctt tttgttgaaa ttttttaaac ctttgtttat
     2701 ttttttttct tcattccgta actcttctac cttctttatt tactttctaa aatccaaata
     2761 caaaacataa aaataaataa acacagagta aattcccaaa ttattccatc attaaaagat
     2821 acgaggcgcg tgtaagttac aggcaagcga tccgtccgga tccaaattat gccaccgtta
     2881 ccttacgttt catggttaat catcgtttac tgccgcctat gagcgtaagc taatgttata
     2941 aagaaacaag ctataatatt gttaaatata gttgatcaac agcattgtaa tgattacaag
     3001 agacgaggtg gaatgaacct tatgaaatgc gtattatata taaactgtaa taagagctaa
     3061 gttgaattga aatctacgat acttgatgtt gacattatag cactagttcc caggaaaccc
     3121 tttcgaaaaa cacagcaaaa acaagagtac tgtaaccaat gtaacatctg tacaccaggg
     3181 acccacacat taccaaaatc aaaattattt ttctaatgcc tgttattttt cctatttttc
     3241 ctctggcgcg tgaatagccc gcagagacgc aaacaatttt cctcgcagtt tttcgcttgt
     3301 ttaatgcgta ttttcccaga taggttcaaa cctttcatct gtatcccgta tatttaagat
     3361 ggcgtttgct ttctccgttg atttttttcc ttcttagtga tttttttgca ttaaatccca
     3421 gaacaatcat ccaactaatc aagaatgcct tccagattca ctaagactag aaagcacaga
     3481 ggtcacgtct caggtatgta gttccatttg gaagagggaa tgaaagaacc aagacggtga
     3541 cttttttttt agtgttgtgc aaccaatatg tcgtgtgtat atcatggtac aggagaatgt
     3601 caatcagcta agtgtactca acatatttct ttgtgttttg attgcgaact ttgtattacc
     3661 atctcactgt tgagacggct tatttgaggt aatagctcga gtaaatgtac tcttccatcg
     3721 caaactgagc aaaaagaaag tgtgcatagc ctttgtcata cttctccttt attataccat
     3781 gatattcaga acagtcatac tgtctactca ttttacggct ataaaaggta actttcattt
     3841 agattatgga aagcactaat tatcgctgta tcaattgaaa tggttgtaga gagcgcaatt
     3901 atgaaaaaga gttaccacgt ttcttttgtt tcgataaaat gtccagttga aaacctgttt
     3961 tactaacgat ttaaaaattg tatttcatta caatattttt tttgtacagc cggtaaaggt
     4021 cgtatcggta agcacagaaa gcaccccggt ggtagaggta tggccggtgg tcaacatcac
     4081 cacagaatta acatggataa ataccatcca ggttatttcg gtatggttgg tatgagatac
     4141 ttccacaagc aacaagctca tttctggaag ccagtcttga acttggacaa attgtggaca
     4201 ttgatcccag aagacaagag agaccaatac ttgaaatctg cttctaagga aactgctcca
     4261 gttattgaca ctttggcagc cggttacggt aagatcttgg gtaagggtag aattccaaat
     4321 gttccagtta tcgtcaaagc tagattcgtc tccaagttgg ctgaagaaaa aatcagagct
     4381 gctggtggtg ttgttgaatt gatcgcttaa gcgcatcaac aaaaactcta tgtattttcc
     4441 aataaattat atatcttcag tttaatctaa ttcaacatct acttctgtat tatttctatc
     4501 acccattttc accgtttttt cgtcttgtct attttgaagt taccagtggc agatcccaac
     4561 gcttttatcg cgctagcact tctaaataag cgaatttctt atgatttatg atttttatta
     4621 ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct taggttttaa
     4681 aacgaaaatt cttattcttg agtaactctt tcctgtaggt caggttgctt tctcaggtat
     4741 agtatgaggt cgctcttatt gaccacacct ctaccggcag atccgctagg gataacaggg
     4801 taatatagat ctgtttagct tgcctcgtcc ccgccgggtc acccggccag cgacatggag
     4861 gcccagaata ccctccttga cagtcttgac gtgcgcagct caggggcatg atgtgactgt
     4921 cgcccgtaca tttagcccat acatccccat gtataatcat ttgcatccat acattttgat
     4981 ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc agacctgcga gcagggaaac
     5041 gctcccctca cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag aaatataaaa
     5101 ggttaggatt tgccactgag gttcttcttt catatacttc cttttaaaat cttgctagga
     5161 tacagttctc acatcacatc cgaacataaa caaccatggg taggagggct tttgtagaaa
     5221 gaaatacgaa cgaaacgaaa atcagcgttg ccatcgcttt ggacaaagct cccttacctg
     5281 aagagtcgaa ttttattgat gaacttataa cttccaagca tacaaaccaa aagggagaac
     5341 aagtaatcca agtagacacg ggaattggat tcttggatca catgtatcat gcactggcta
     5401 aacatgcagg ctggagctta cgactttact caagaggtga tttaatcatc gatgatcatc
     5461 acactgcaga agatactgct attgcacttg gtattgcatt caagcaggct atgagtaact
     5521 ttgccggcgt taaaagattt ggacatgctt attgtccact tgacgaagct ctttctagaa
     5581 gcgtagttga cttgtcggga cggccctatg ctgttatcga tttgggatta aagcgtgaaa
     5641 aggttgggga attgtcctgt gaaatgatcc ctcacttact atattccttt tcggtagcag
     5701 ctggaattac tttgcatgtt acctgcttat atggtagtaa tgaccatcat cgtgctgaaa
     5761 gcgcttttaa atctctggct gttgccatgc gcgcggctac tagtcttact ggaagttctg
     5821 aagtcccaag cacgaaggga gtgttgtaaa gagtactgac aataaaaaga ttcttgtttt
     5881 caagaacttg tcatttgtat agttttttta tattgtagtt gttctatttt aatcaaatgt
     5941 tagcgtgatt tatatttttt ttcgcctcga catcatctgc ccagatgcga agttaagtgc
     6001 gcagaaagta atatcatgcg tcaatcgtat gtgaatgctg gtcgctatac tgctgtcgat
     6061 tcgatactaa cgccgccatc cagtttaaac gagctcgaat tcatcgatga tatcagatcc
     6121 actagtggcc tatgcggccg cggatctgcc ggtctcccta tagtgagtcg tattaatttc
     6181 gataagccag gttaacctgc attaatgaat cggccaac
//
