LOCUS       sequence-263316-        5712 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses N-terminal 6xHis-tagged S9 ribosomal protein in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5712)
  AUTHORS   Duss O, Stepanyuk GA, Puglisi JD, Williamson JR
  TITLE     Transient Protein-RNA Interactions Guide Nascent Ribosomal RNA 
            Folding.
  JOURNAL   Cell. 2019 Nov 27;179(6):1357-1369.e16. doi: 
            10.1016/j.cell.2019.10.035. Epub 2019 Nov 21.
  PUBMED    31761533
REFERENCE   2  (bases 1 to 5712)
  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: 
            "10.1016/j.cell.2019.10.035"; journalName: "Cell"; date: 
            "2019-11-27- 27"; volume: "179"; issue: "6"; pages: "1357-1369.e16"
COMMENT     Sequence Label: pET-S9
FEATURES             Location/Qualifiers
     source          1..5712
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     102..121
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     205..224
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        205..223
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    224..248
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             263..285
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             305..322
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             781..798
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(851..869)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      865..912
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      949..1404
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1036..1055)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1246..1267
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1496..2311)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1496..2311)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2219..2238
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2303..2322
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2433..3021
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2922..2941
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3175..3192
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3207..3346
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3272..3291)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3432..3454
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3448..3639)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4448..5530)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5492..5511
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5531..5608)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa
       61 gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag
      121 gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg
      181 atcgagatct cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata
      241 acaattcccc tctagaaata attttgttta actttaagaa ggagatatac atatgggcag
      301 cagccatcat catcatcatc actggggcgc tgaaaatcaa tactacggca ctggtcgccg
      361 caaaagttcc gcagctcgcg ttttcatcaa accgggcaac ggtaaaatcg taatcaacca
      421 acgttctctg gaacagtact tcggtcgtga aactgcccgc atggtagttc gtcagccgct
      481 gtgtctggtc gacatggttg agaaactgga cctgtacatc accgttaaag gtggtggtat
      541 ctctggtcag gctggtgcga tccgtcacgg tatcacccgc gctctgatgg aatacgacga
      601 gtccctgcgt tctgaactgc gtaaagctgg cttcgttact cgtgacgctc gtcaggttga
      661 acgtaagaaa gtcggtctgc gtaaagcacg tcgtcgtccg cagttctcca aacgttaaag
      721 acaagcttag gtatttattc ggcgcaaagt gcgtcgggtg atgctgccaa cttagtcgag
      781 caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg
      841 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
      901 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
      961 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     1021 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     1081 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     1141 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     1201 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     1261 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     1321 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     1381 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     1441 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga
     1501 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     1561 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     1621 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     1681 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     1741 ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat
     1801 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     1861 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     1921 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     1981 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     2041 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     2101 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     2161 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     2221 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag
     2281 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     2341 cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     2401 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     2461 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     2521 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     2581 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     2641 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     2701 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     2761 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     2821 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     2881 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     2941 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3001 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3061 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     3121 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     3181 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     3241 ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     3301 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     3361 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     3421 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     3481 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg
     3541 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa
     3601 gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg
     3661 atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt
     3721 tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg
     3781 gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga
     3841 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     3901 gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca
     3961 tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat
     4021 cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga
     4081 caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc
     4141 gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc
     4201 gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa
     4261 aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat
     4321 aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     4381 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     4441 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4501 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     4561 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     4621 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     4681 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa
     4741 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     4801 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     4861 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     4921 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     4981 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     5041 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     5101 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     5161 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     5221 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     5281 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     5341 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     5401 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     5461 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     5521 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     5581 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     5641 tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat
     5701 ggtgcatgca ag
//
