LOCUS       sequence-457025-        4988 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses RssB with N256A mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4988)
  AUTHORS   Dorich V, Brugger C, Tripathi A, Hoskins JR, Tong S, Suhanovsky MM, 
            Sastry A, Wickner S, Gottesman S, Deaconescu AM
  TITLE     Structural basis for inhibition of a response regulator of sigma(S) 
            stability by a ClpXP antiadaptor.
  JOURNAL   Genes Dev. 2019 Jun 1;33(11-12):718-732. doi: 
            10.1101/gad.320168.118. Epub 2019 Apr 11.
  PUBMED    30975721
REFERENCE   2  (bases 1 to 4988)
  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: "Genes 
            Dev."; date: "2019-06-1"; volume: "33(11-12):718-732. doi"; pages: "
            10.1101/gad.320168.118. Epub 2019 Apr 11"
COMMENT     Sequence Label: pVF29
FEATURES             Location/Qualifiers
     source          1..4988
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     6..25
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(244..348)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     416..434
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     637..656
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        637..655
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    656..680
                     /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             695..717
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1785..1802
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1855..1873)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1869..1916
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1953..2408
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2040..2059)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2250..2271
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2977..2996
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3058..3077
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(3527..3544)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3661..4206)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(3741..3760)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4371..4988,1..174,175..243))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
       61 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
      121 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
      181 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
      241 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
      301 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
      361 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
      421 gcgtatcacg aggccctttc gtcttcaaga attttataaa ccgtggagcg ggcaatactg
      481 agctgatgag caatttccgt tgcaccagtg cccttctgat gaagcgtcag cacgacgttc
      541 ctgtccacgg tacgcctgcg gccaaatttg attcctttca gctttgcttc ctgtcggccc
      601 tcattcgtgc gctctaggat ctcgatcccg cgaaattaat acgactcact ataggggaat
      661 tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat
      721 acatatgacg cagccattgg tcggaaaaca gattctcatt gttgaagatg agcaggtatt
      781 tcgctcgctt ctggattcat ggttttcctc attgggagcg acaacggtac tggcggctga
      841 tggggtggat gcccttgagt tgctgggagg tttcactcca gacctgatga tatgtgatat
      901 cgcgatgcca cgaatgaacg ggcttaaact gctggagcat atacgtaaca gaggcgacca
      961 gaccccagtt ctggtgatat ctgccactga aaatatggca gatattgcca aagcgttacg
     1021 tctgggcgtt gaagatgttt tgctgaaacc agttaaagat ctgaatcgct tgcgcgagat
     1081 ggtttttgcc tgtctctatc ccagcatgtt taattcgcgc gttgaggaag aggaaaggct
     1141 ttttcgcgac tgggatgcaa tggttgataa ccctgccgca gcggcgaaat tattacagga
     1201 actacaaccg ccggttcagc aggtgatttc ccattgccgg gttaattatc gtcaattggt
     1261 tgccgcggac aaacccggcc tggtgcttga tattgccgca ctttcggaaa acgatctggc
     1321 attttattgc cttgatgtca cccgagctgg acataatggc gtacttgctg ccttgttatt
     1381 acgcgcattg tttaacggat tattacagga acagcttgca caccaaaatc aacggttgcc
     1441 agagttgggc gcgttattga agcaggtaaa ccatttactt cgtcaggccg ctctgccggg
     1501 gcagtttccg ctattagttg gctattatca tcgcgaactg aaaaatctca ttctggtttc
     1561 tgcgggtctg aatgcgacgt taaataccgg cgaacaccag gtgcaaatca gtaatggtgt
     1621 tccgttaggc actttaggta acgcttattt gaatcaattg agccagcgat gcgatgcctg
     1681 gcaatgccaa atatggggaa ccggtggtcg actgcgcttg atgttgtctg cagaatgagg
     1741 atccgaattc gagctccgtc gacaagcttg cggccgcact cgagcaccac caccaccacc
     1801 actgagatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg
     1861 agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga
     1921 aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
     1981 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     2041 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2101 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2161 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     2221 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     2281 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     2341 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     2401 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     2461 taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat
     2521 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
     2581 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
     2641 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
     2701 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
     2761 cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat
     2821 ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc
     2881 attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata
     2941 caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata
     3001 taaatcagca tccatgttgg aatttaatcg cggcagataa gacgtttccc gttgaatatg
     3061 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgatga
     3121 tatattttta tcttgtgcaa tgtaacatca gagattttga gacacaacgt ggctttgttg
     3181 aataaatcga acttttgctg agttgaagga tcagatcacg catcttcccg acaacgcaga
     3241 ccgttccgtg gcaaagcaaa agttcaaaat caccaactgg tccacctaca acaaagctct
     3301 catcaaccgt ggctccctca ctttctggct ggatgatggg gcgattcagg cctggaatga
     3361 gtcagcaaca ccttcttcac gaggcagacc tcagcgctag cggagtgtat actggcttac
     3421 tatgttggca ctgatgaggg tgtcagtgaa gtgcttcatg tggcaggaga aaaaaggctg
     3481 caccggtgcg tcagcagaat atgtgataca ggatatattc cgcttcctcg ctcactgact
     3541 cgctacgctc ggtcgttcga ctgcggcgag cggaaatggc ttacgaacgg ggcggagatt
     3601 tcctggaaga tgccaggaag atacttaaca gggaagtgag agggccgcgg caaagccgtt
     3661 tttccatagg ctccgccccc ctgacaagca tcacgaaatc tgacgctcaa atcagtggtg
     3721 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggcggct ccctcgtgcg
     3781 ctctcctgtt cctgcctttc ggtttaccgg tgtcattccg ctgttatggc cgcgtttgtc
     3841 tcattccacg cctgacactc agttccgggt aggcagttcg ctccaagctg gactgtatgc
     3901 acgaaccccc cgttcagtcc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3961 acccggaaag acatgcaaaa gcaccactgg cagcagccac tggtaattga tttagaggag
     4021 ttagtcttga agtcatgagc cggttaaggc taaactgaaa ggacaagttt tggtgactgc
     4081 gctcctccaa gccagttacc tcggttcaaa gagttggtag ctcagagaac cttcgaaaaa
     4141 ccgccctgca aggcggtttt ttcgttttca gagcaagaga ttacgcgcag accaaaacga
     4201 tctcaagaag atcatcttat taaggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4261 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4321 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4381 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4441 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     4501 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     4561 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     4621 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     4681 attgctgcag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     4741 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     4801 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     4861 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     4921 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     4981 gcgtcaac
//
