LOCUS       sequence-223545-        6685 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses C-terminus of E. coli NusG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6685)
  AUTHORS   Lawson MR, Ma W, Bellecourt MJ, Artsimovitch I, Martin A, Landick R,
            Schulten K, Berger JM
  TITLE     Mechanism for the Regulated Control of Bacterial Transcription 
            Termination by a Universal Adaptor Protein.
  JOURNAL   Mol Cell. 2018 Sep 20;71(6):911-922.e4. doi: 
            10.1016/j.molcel.2018.07.014. Epub 2018 Aug 16.
  PUBMED    30122535
REFERENCE   2  (bases 1 to 6685)
  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: 
            "10.1016/j.molcel.2018.07.014"; journalName: "Mol Cell"; date: 
            "2018-09-20- 20"; volume: "71"; issue: "6"; pages: "911-922.e4"
COMMENT     Sequence Label: EcNusGKOW
FEATURES             Location/Qualifiers
     source          1..6685
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             15..37
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             56..73
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             83..1183
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1157..1180
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1238..1258
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1505..1522
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1575..1593)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1589..1636
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1673..2128
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1760..1779)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1970..1991
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2221..3036)
                     /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(2221..3036)
                     /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     2944..2963
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3028..3047
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3158..3746
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3647..3666
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3900..3917
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3932..4071
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3997..4016)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4157..4179
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4173..4364)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5173..6255)
                     /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     6217..6236
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6256..6333)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6539..6558
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6642..6661
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6642..6660
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6661..6685
                     /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)."
ORIGIN
        1 cctctagaaa taattttgtt taactttaag aaggagatat accatgggtt cttctcacca
       61 tcaccatcac catggttctt ctatgaaaat cgaagaaggt aaactggtaa tctggattaa
      121 cggcgataaa ggctataacg gtctcgctga agtcggtaag aaattcgaga aagataccgg
      181 aattaaagtc accgttgagc atccggataa actggaagag aaattcccac aggttgcggc
      241 aactggcgat ggccctgaca ttatcttctg ggcacacgac cgctttggtg gctacgctca
      301 atctggcctg ttggctgaaa tcaccccgga caaagcgttc caggacaagc tgtatccgtt
      361 tacctgggat gccgtacgtt acaacggcaa gctgattgct tacccgatcg ctgttgaagc
      421 gttatcgctg atttataaca aagatctgct gccgaacccg ccaaaaacct gggaagagat
      481 cccggcgctg gataaagaac tgaaagcgaa aggtaagagc gcgctgatgt tcaacctgca
      541 agaaccgtac ttcacctggc cgctgattgc tgctgacggg ggttatgcgt tcaagtatga
      601 aaacggcaag tacgacatta aagacgtggg cgtggataac gctggcgcga aagcgggtct
      661 gaccttcctg gttgacctga ttaaaaacaa acacatgaat gcagacaccg attactccat
      721 cgcagaagct gcctttaata aaggcgaaac agcgatgacc atcaacggcc cgtgggcatg
      781 gtccaacatc gacaccagca aagtgaatta tggtgtaacg gtactgccga ccttcaaggg
      841 tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt attaacgccg ccagtccgaa
      901 caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg actgatgaag gtctggaagc
      961 ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag tcttacgagg aagagttggc
     1021 gaaagatcca cgtattgccg ccactatgga aaacgcccag aaaggtgaaa tcatgccgaa
     1081 catcccgcag atgtccgctt tctggtatgc cgtgcgtact gcggtgatca acgccgccag
     1141 cggtcgtcag actgtcgatg aagccctgaa agacgcgcag actaattcga gctcgaacaa
     1201 caacaacaat aacaataaca acaacctcgg gatcgaggaa aacctgtact tccaatccaa
     1261 tgcaaaaacg ctgtttgaac cgggtgaaat ggtccgtgtt aatgatggtc cgttcgctga
     1321 cttcaacggt gttgttgaag aagtggatta cgagaaatct cgtctgaaag tgtctgtttc
     1381 tatcttcggt cgtgcgaccc cggtagagct ggacttcagc caggttgaaa aagcctaata
     1441 acattggaag tggataacgg atccgaattc gagcgccgtc gacaagcttg cggccgcact
     1501 cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa aggaagctga
     1561 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     1621 cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg ggacgcgccc
     1681 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     1741 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     1801 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     1861 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     1921 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     1981 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2041 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2101 tttaacaaac tagtaacgtt tacaatttca ggtggcactt ttcggggaaa tgtgcgcgga
     2161 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gaattaattc
     2221 ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat
     2281 accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca
     2341 taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc
     2401 tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac
     2461 tgaatccggt gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca
     2521 gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg
     2581 cgcctgagcg agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga
     2641 atgcaaccgg cgcaggaaca ctgccagcgc atcaacaatg ttttcacctg aatcaggata
     2701 ttcttctaat acctggaatg ctgttttccc ggggatcgca gtggtgagta accatgcatc
     2761 atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt
     2821 tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa
     2881 caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac
     2941 attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg
     3001 cctagagcaa gacgtttccc gttgaatatg gctcataaca ccccttgtat tactgtttat
     3061 gtaagcagac agttttattg ttcatgacca aaatccctta acgtgagttt tcgttccact
     3121 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3181 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3241 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3301 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3361 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3421 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3481 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     3541 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     3601 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     3661 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     3721 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     3781 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     3841 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     3901 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     3961 tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata
     4021 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     4081 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     4141 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     4201 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct
     4261 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     4321 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     4381 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     4441 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     4501 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat
     4561 acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata
     4621 atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc
     4681 attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg
     4741 cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc
     4801 aacgacagga gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc
     4861 ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag
     4921 attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg
     4981 ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca
     5041 gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg
     5101 aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta
     5161 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5221 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct
     5281 tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg
     5341 cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa
     5401 cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc
     5461 accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt
     5521 ggcaaccagc atcgcagtgg gaacgatacc ctcattcagc atttgcatgg tttgttgaaa
     5581 accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt
     5641 gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc
     5701 taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc
     5761 gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa
     5821 cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata
     5881 gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc
     5941 ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg
     6001 agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac
     6061 gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt
     6121 cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg
     6181 gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa
     6241 cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat
     6301 accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg
     6361 actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa
     6421 ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca
     6481 tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg
     6541 tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga
     6601 tgcgtccggc gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg
     6661 ggaattgtga gcggataaca attcc
//
