LOCUS       sequence-446215-        8158 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsNot6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8158)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 8158)
  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"
COMMENT     Sequence Label: pETM-41-HsNot6_N
FEATURES             Location/Qualifiers
     source          1..8158
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             94..111
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             115..1215
                     /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     1189..1212
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1267..1287
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3023..3040
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3093..3111)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3107..3154
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3191..3646
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3278..3297)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3488..3509
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3738..4553)
                     /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(3738..4553)
                     /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     4461..4480
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4545..4564
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4675..5263
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5164..5183
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5417..5434
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5449..5588
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5514..5533)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5674..5696
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5690..5881)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6690..7772)
                     /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     7734..7753
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7773..7850)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     8056..8075
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg aaacatcacc atcaccatca ccccatgaaa
      121 atcgaagaag gtaaactggt aatctggatt aacggcgata aaggctataa cggtctcgct
      181 gaagtcggta agaaattcga gaaagatacc ggaattaaag tcaccgttga gcatccggat
      241 aaactggaag agaaattccc acaggttgcg gcaactggcg atggccctga cattatcttc
      301 tgggcacacg accgctttgg tggctacgct caatctggcc tgttggctga aatcaccccg
      361 gacaaagcgt tccaggacaa gctgtatccg tttacctggg atgccgtacg ttacaacggc
      421 aagctgattg cttacccgat cgctgttgaa gcgttatcgc tgatttataa caaagatctg
      481 ctgccgaacc cgccaaaaac ctgggaagag atcccggcgc tggataaaga actgaaagcg
      541 aaaggtaaga gcgcgctgat gttcaacctg caagaaccgt acttcacctg gccgctgatt
      601 gctgctgacg ggggttatgc gttcaagtat gaaaacggca agtacgacat taaagacgtg
      661 ggcgtggata acgctggcgc gaaagcgggt ctgaccttcc tggttgacct gattaaaaac
      721 aaacacatga atgcagacac cgattactcc atcgcagaag ctgcctttaa taaaggcgaa
      781 acagcgatga ccatcaacgg cccgtgggca tggtccaaca tcgacaccag caaagtgaat
      841 tatggtgtaa cggtactgcc gaccttcaag ggtcaaccat ccaaaccgtt cgttggcgtg
      901 ctgagcgcag gtattaacgc cgccagtccg aacaaagagc tggcaaaaga gttcctcgaa
      961 aactatctgc tgactgatga aggtctggaa gcggttaata aagacaaacc gctgggtgcc
     1021 gtagcgctga agtcttacga ggaagagttg gcgaaagatc cacgtattgc cgccactatg
     1081 gaaaacgccc agaaaggtga aatcatgccg aacatcccgc agatgtccgc tttctggtat
     1141 gccgtgcgta ctgcggtgat caacgccgcc agcggtcgtc agactgtcga tgaagccctg
     1201 aaagacgcgc agactaattc gagctcgaac aacaacaaca ataacaataa caacaacccc
     1261 atgagcgaga atctttattt tcagggcgcc atggacggta ccatgcccaa agaaaaatac
     1321 gagccccctg accctcggag gatgtataca attatgtctt ctgaggaagc agcaaatgga
     1381 aagaaatccc actgggcaga gcttgaaata agtggaaaag taagaagctt aagcgcatct
     1441 ttgtggtcac taactcacct gacagctttg catttgagtg acaattccct gtcccgaatt
     1501 ccttcagaca ttgccaagct tcacaatctg gtgtatttgg acctgtcatc taataaaatt
     1561 cgtagcttac ccgcagaact cggaaacatg gtatcactca gggagctcca tttaaataac
     1621 aacctgttac gagttctacc ttttgagctg ggaaaactgt ttcagttgca gactttaggc
     1681 ctgaaaggaa atccccttac ccaggatata ttgaaccttt atcaggaacc agatggaaca
     1741 agacggctgc tgaactattt gcttgataat ttgtcaggta ctgcaaaaag aattacaaca
     1801 gaacaaccac ctccaaggtc ttggattatg ttacaagaac cagataggac aaggccaact
     1861 gccttgtttt ctgtcatgtg ctataatgtt ctttgtgata aatatgcgac ccggcagtta
     1921 tacggctact gtccatcatg ggcgctaaac tgggactaca ggaaaaaggc cattattcaa
     1981 gaaatcttga gctgcaatgc tgatatcgta agtcttcagg aggttgaaac ggaacagtat
     2041 tacagttttt ttctggtaga gctgaaagaa cgtggctata atggattctt cagtcctaag
     2101 tctagagcta ggacaatgtc agaacaagaa aggaaacatg ttgatggctg tgcaatattc
     2161 ttcaagacag aaaaatttac tttggttcag aaacacactg ttgaatttaa tcagctagcc
     2221 atggcaaatt ctgaggggtc tgaagctatg ctgaacagag tcatgacaaa agataacatt
     2281 ggggttgcag tactgctaga acttcggaag gaatcgattg aaatgccgtc tggaaagcca
     2341 catcttggaa cagaaaaaca acttattctt gtggctaacg cccacatgca ttgggaccct
     2401 gaatactctg atgtgaagtt ggtacaaact atgatgttcc tctcagaagt gaagaacatt
     2461 attgataaag cctctcgcaa cctcaaatcc agtgttttgg gagaatttgg aactattcca
     2521 cttgtgttat gtgcagatct taattctttg ccagactctg gtgttgtaga atatttgagc
     2581 acaggtggag tagaaacaaa tcacaaagac tttaaggagt tgaggtataa tgaaagtctc
     2641 acaaacttca gctgtcatgg gaagaatgga accaccaatg gaaggatcac tcatggtttc
     2701 aagttacaga gtgcctatga gagtggcctg atgccttaca cgaattacac atttgatttc
     2761 aagggtataa tagactacat tttctattct aaacctcagc tgaacacctt aggcatcctg
     2821 ggccctctgg accaccactg gctggttgag aataacatca gtggctgccc gcaccccctc
     2881 atcccctctg accacttctc actttttgca caactggagc tcttactgcc tttcctgccc
     2941 caagtcaacg gcatccacct tcctggcagg aggtagggat ccgaattcga gctccgtcga
     3001 caagcttgcg gccgcactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa
     3061 agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag cataacccct
     3121 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatt
     3181 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3241 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     3301 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3361 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     3421 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3481 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     3541 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3601 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     3661 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     3721 cgctcatgaa ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc
     3781 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     3841 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     3901 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     3961 tcaccatgag tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag
     4021 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg
     4081 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa
     4141 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     4201 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg
     4261 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     4321 aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct
     4381 ttgccatgtt tcagaaacaa ctctggcgca tcgggctttc catacaatcg atagattgtc
     4441 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     4501 ttggaattta atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc
     4561 cttgtattac tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg
     4621 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     4681 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     4741 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     4801 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     4861 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4921 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4981 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     5041 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     5101 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     5161 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     5221 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     5281 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     5341 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     5401 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     5461 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     5521 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     5581 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     5641 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     5701 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     5761 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     5821 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     5881 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     5941 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     6001 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     6061 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     6121 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     6181 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     6241 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     6301 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     6361 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     6421 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     6481 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     6541 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     6601 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     6661 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     6721 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     6781 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     6841 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     6901 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     6961 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     7021 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     7081 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     7141 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     7201 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     7261 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     7321 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     7381 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     7441 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     7501 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     7561 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     7621 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     7681 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     7741 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     7801 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     7861 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     7921 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     7981 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     8041 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     8101 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaat
//
