LOCUS       sequence-350250-        5710 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Rok _Linker overexpression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5710)
  AUTHORS   Erkelens AM, Qin L, van Erp B, Miguel-Arribas A, Abia D, Keek HGJ, 
            Markus D, Cajili MKM, Schwab S, Meijer WJJ, Dame RT
  TITLE     The B. subtilis Rok protein is an atypical H-NS-like protein 
            irresponsive to physico-chemical cues.
  JOURNAL   Nucleic Acids Res. 2022 Nov 21. pii: 6835367. doi: 
            10.1093/nar/gkac1064.
  PUBMED    36408910
REFERENCE   2  (bases 1 to 5710)
  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: "Nucleic 
            Acids Res. 2022 Nov 21. pii: 6835367. doi: 10.1093/nar/gkac1064."
COMMENT     Sequence Label: pRD415
FEATURES             Location/Qualifiers
     source          1..5710
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      75..530
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(162..181)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     372..393
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(622..1437)
                     /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(622..1437)
                     /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     1345..1364
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1429..1448
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1559..2147
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2048..2067
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2301..2318
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2333..2472
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2398..2417)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2558..2580
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2574..2765)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3574..4656)
                     /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
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTAPPR
                     ALADSLMQLVRQVSRLESGQ"
     primer_bind     4618..4637
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4657..4734)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4940..4959
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5047..5066
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5047..5065
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5066..5090
                     /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             5105..5127
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(5687..5705)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      join(5701..5710,1..38)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
       61 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
      121 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
      181 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
      241 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
      301 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
      361 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
      421 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
      481 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact
      541 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
      601 tatccgctca tgaattaatt cttagaaaaa ctcatcgagc atcaaatgaa actgcaattt
      661 attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga
      721 aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac
      781 tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga
      841 gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt
      901 ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa
      961 accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg
     1021 acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat
     1081 attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc cggggatcgc
     1141 agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg
     1201 cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct
     1261 acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat
     1321 tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc
     1381 catgttggaa tttaatcgcg gcctagagca agacgtttcc cgttgaatat ggctcataac
     1441 accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgacc aaaatccctt
     1501 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     1561 gaaatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     1621 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     1681 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     1741 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     1801 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     1861 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     1921 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     1981 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     2041 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     2101 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     2161 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     2221 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     2281 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc
     2341 ggtattttct ccttacgcat ctgtgcggta tttcacaccg caatggtgca ctctcagtac
     2401 aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg
     2461 gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     2521 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     2581 ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg
     2641 tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag tttctccaga
     2701 agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg
     2761 gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa
     2821 cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt
     2881 tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt
     2941 caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct
     3001 gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac
     3061 gaaacacgga aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag
     3121 cagtcgcttc acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc
     3181 cgccagccta gccgggtcct caacgacagg agcacgatca tgcgcacccg tggggccgcc
     3241 atgccggcga taatggcctg cttctcgccg aaacgtttgg tggcgggacc agtgacgaag
     3301 gcttgagcga gggcgtgcaa gattccgaat accgcaagcg acaggccgat catcgtcgcg
     3361 ctccagcgaa agcggtcctc gccgaaaatg acccagagcg ctgccggcac ctgtcctacg
     3421 agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac
     3481 cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa
     3541 tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     3601 ctgtcgtacc agctgcatta atgaatcggc caacgcgcgg ggaggggcgg tttgcgtatt
     3661 gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac
     3721 cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa
     3781 atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta
     3841 tcccactacc gagatgtccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc
     3901 gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag
     3961 catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat
     4021 cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga
     4081 gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg
     4141 ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg
     4201 gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc
     4261 atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt
     4321 gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct
     4381 ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgtag
     4441 ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc
     4501 cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt
     4561 cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc
     4621 atactctgcg acatcgtata acgttactgg tttcacattc accaccctga attgactctc
     4681 ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat
     4741 ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt aggttgaggc
     4801 cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc aacagtcccc
     4861 cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc ccgaagtggc
     4921 gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg
     4981 cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatcgat ctcgatcccg
     5041 cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa
     5101 taattttgtt taactttaag aaggagatat acatatgttt aatgaaagag aagctttgcg
     5161 cttgcggtta gaacaattaa atgaagctga agtgaaagtc attcgtgaat atcaaattga
     5221 acgtgataaa atatacgcaa aattaagaga gctggacaga aatggaagcc cttccgaaat
     5281 caaaaaggat ttccgttctg aaaaaaaacc tgactctctg ccggttctcg ctgagcttgc
     5341 ggctcaggaa atcagagccg gtataccaga cggaacaaca cgaagaagaa gaggaacggc
     5401 aagaccggga tcaaaagcag ctaagctgcg tgaagctgct attaaaacat taaaacgcca
     5461 caacgcggcg attaagagct cggagcttca aaaagagatt gaaaaagaaa gcggtcttga
     5521 aatccctaat atgacaacgt ttatgcaaag cttaattaaa atgtacccgg aagtgaaaaa
     5581 gccatatcgc gggcagtaca ttttagaggg tgaaatcgag tcggcggaat cagcaaacga
     5641 ataactgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa
     5701 ctagcataac
//
