LOCUS       sequence-291120-        7518 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsNot1_4-682.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7518)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 7518)
  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: pnYC-NpM-HsNot1_4-682_AF
FEATURES             Location/Qualifiers
     source          1..7518
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2294..2313
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2402..2421
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2402..2420
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2421..2445
                     /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             2460..2482
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2489..3589
                     /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="METEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     CDS             3605..3628
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(5750..5768)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5764..5811
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(5986..5991,5992..6008,6009..6014))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     rep_origin      6126..6671
                     /direction=RIGHT
                     /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     6572..6591
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6788..6805
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        7197..7299
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             join(7300..7518,1..441)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFGMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(7366..7385)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 ggtatggcaa tgaaagacgg tgagctggtg atatgggata gtgttcaccc ttgttacacc
       61 gttttccatg agcaaactga aacgttttca tcgctctgga gtgaatacca cgacgatttc
      121 cggcagtttc tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat
      181 ttccctaaag ggtttattga gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc
      241 accagttttg atttaaacgt ggccaatatg gacaacttct tcgcccccgt tttcaccatg
      301 ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc tggcgattca ggttcatcat
      361 gccgtctgtg atggcttcca tgtcggcaga atgcttaatg aattacaaca gtactgcgat
      421 gagtggcagg gcggggcgta atttttttaa ggcagttatt ggtgccctta aacgcctggt
      481 gctacgcctg aataagtgat aataagcgga tgaatggcag aaattcgaaa gcaaattcga
      541 cccggtcgtc ggttcagggc agggtcgtta aatagccgct tatgtctatt gctggtttac
      601 cggtttattg actaccggaa gcagtgtgac cgtgtgcttc tcaaatgcct gaggccagtt
      661 tgctcaggct ctccccgtgg aggtaataat tgacgatatg atcatttatt ctgcctccca
      721 gagcctgata aaaacggtta gcgcttcgtt aatacagatg taggtgttcc acagggtagc
      781 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgcttg tttcggcgtg
      841 ggtatggtgg caggccccgt ggccggggga ctgttgggcg ctgccggcac ctgtcctacg
      901 agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac
      961 cggaaggagc taccggacag cggtgcggac tgttgtaact cagaataaga aatgaggccg
     1021 ctcatggcgt tgactctcag tcatagtatc gtggtatcac cggttggttc cactctctgt
     1081 tgcgggcaac ttcagcagca cgtaggggac ttccgcgttt ccagacttta cgaaacacgg
     1141 aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt
     1201 cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct
     1261 agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg
     1321 cccgagatgc gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa
     1381 gggttggttt gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg
     1441 gtgaatccgt tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg
     1501 caccgcgacg caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca
     1561 acccgttcca tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccagtga
     1621 tcgaagttag gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat
     1681 ctacctgcct ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa
     1741 gaatcataat ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca
     1801 gcgcgtcggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     1861 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     1921 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     1981 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     2041 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgac
     2101 gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga
     2161 gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca
     2221 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
     2281 gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg
     2341 tgatgccggc cacgatgcgt ccggcgtaga ggatcagact aagatctcga tcccgcgaaa
     2401 ttaatacgac tcactatagg ggaattgtga gcggataaca attccccact agtaataatt
     2461 ttgtttaact ttaagaagga gatataccat ggaaactgaa gaaggtaaac tggtaatctg
     2521 gattaacggc gataaaggct ataacggtct cgctgaagtc ggtaagaaat tcgagaaaga
     2581 taccggaatt aaagtcaccg ttgagcatcc ggataaactg gaagagaaat tcccacaggt
     2641 tgcggcaact ggcgatggcc ctgacattat cttctgggca cacgaccgct ttggtggcta
     2701 cgctcaatct ggcctgttgg ctgaaatcac cccggacaaa gcgttccagg acaagctgta
     2761 tccgtttacc tgggatgccg tacgttacaa cggcaagctg attgcttacc cgatcgctgt
     2821 tgaagcgtta tcgctgattt ataacaaaga cctgctgccg aacccgccaa aaacctggga
     2881 agagatcccg gcgctggata aagaactgaa agcgaaaggt aagagcgccc tgatgttcaa
     2941 cctgcaagaa ccgtacttca cctggccgct gattgctgct gacgggggtt atgcgttcaa
     3001 gtatgaaaac ggcaagtacg acattaaaga cgtgggcgtg gataacgctg gcgcgaaagc
     3061 gggtctgacc ttcctggttg acctgattaa aaacaaacac atgaatgcag acaccgatta
     3121 ctccatcgca gaagctgcct ttaataaagg cgaaacagcg atgaccatca acggcccgtg
     3181 ggcatggtcc aacatcgaca ccagcaaagt gaattatggt gtaacggtac tgccgacctt
     3241 caagggtcaa ccatccaaac cgttcgttgg cgtgctgagc gcaggtatta acgccgccag
     3301 tccgaacaaa gagctggcaa aagagttcct cgaaaactat ctgctgactg atgaaggtct
     3361 ggaagcggtt aataaagaca aaccgctggg tgccgtagcg ctgaagtctt acgaggaaga
     3421 gttggcgaaa gatccacgta ttgccgccac gatggaaaac gcccagaaag gtgaaatcat
     3481 gccgaacatc ccgcagatgt ccgctttctg gtatgccgtg cgtactgcgg tgatcaacgc
     3541 cgccagcggt cgtcagactg tcgatgaagc ccttaaagac gcgcagactg gtaccggcag
     3601 cggcctggaa gttctgttcc aggggcccca tatgctcgag gactcgctct cgctggcctt
     3661 gtctcaaatc agctacctgg tggacaattt aaccaagaaa aattaccgag ccagccagca
     3721 ggaaatacag catattgtga atcggcacgg tcctgaggca gacaggcatt tattacgctg
     3781 cctattttcg catgtggatt tcagtggcga tggtaaaagc agtggcaaag atttccatca
     3841 gactcagttt ctgattcagg agtgtgcgtt gctgattaca aagccaaatt ttatctcgac
     3901 gctgtcctat gccattgata atccattgca ctatcagaag agtttaaagc ctgcacccca
     3961 cttatttgcc cagctgagta aagtgctcaa attaagcaaa gtacaagagg taatttttgg
     4021 ccttgccctg ttgaattctt ccagctcaga tcttagaggt ttcgctgccc agtttatcaa
     4081 acagaagctt ccagatcttc tgcgttctta cattgacgca gacgtcagtg gaaatcaaga
     4141 aggtggcttc caagatatag caatagaggt cctacacctc ctcctctccc atctcctctt
     4201 tgggcagaag ggagcctttg gagttggaca agaacagata gacgcttttc ttaagacgct
     4261 gcgcagagat tttccccaag aacgctgtcc cgtggtgctc gcaccacttt tataccctga
     4321 aaaacgggac attctaatgg acaggatcct gcctgattcc ggaggggtag ctaaaaccat
     4381 gatggagagc tctttggctg atttcatgca agaagtaggc tatggctttt gtgcaagtat
     4441 tgaagaatgt cgcaatataa tcgtgcagtt tggtgttcgg gaggtcacag ctgcccaggt
     4501 tgcaagggtt ttgggaatga tggctcgaac tcattcagga ttaacagatg gcattccatt
     4561 acagagtatt tctgctccgg gcagtgggat ctggagtgat gggaaagata aaagtgatgg
     4621 agcacaggca cacacatgga atgtagaagt cttgattgac gttcttaaag aactgaatcc
     4681 aagtttgaat ttcaaggaag taacttatga actggaccat cctggatttc aaattcgtga
     4741 cagtaaagga cttcataatg tggtttatgg cattcagagg ggtttgggta tggaagtgtt
     4801 cccagtagac ctcatatata gaccttggaa acatgctgaa ggccagctct ccttcattca
     4861 acattccctt ataaatccag agatcttctg ttttgctgac tatccctgtc atactgttgc
     4921 cactgatatt ctgaaagcac caccagagga tgacaatcga gaaattgcca catggaagag
     4981 cttggatttg attgaatctc tgctgaggct tgcagaggtt gggcagtatg agcaagtcaa
     5041 acagctcttc agcttcccta tcaaacactg tccagacatg ctggtattgg ccttactaca
     5101 aattaacacc tcttggcata ccttgcgcca tgaacttatc tccactctga tgccaatttt
     5161 ccttggaaac catcctaact cagctattat tttgcactat gcatggcatg ggcagggaca
     5221 gtctccctca attcgccaac ttatcatgca tgcaatggca gaatggtaca tgagagggga
     5281 gcagtatgat caggccaaat tgtctcgaat acttgatgtg gcccaggact tgaaggcctt
     5341 gtcaatgctg ctaaatggta ctccatttgc ctttgttatt gaccttgctg cacttgcttc
     5401 acgtcgtgaa tacctcaaac ttgataagtg gctcacagat aaaattcgag agcatgggga
     5461 gccttttatc caggcgtgta tgactttttt aaagagacgg tgtccttcta ttttgggcgg
     5521 acttgcccca gaaaaagacc agcccaaaag tgctcaactt cctccagaaa ctttggcgac
     5581 aatgttggcc tgtctgcaag cttgtgcagg gagtgtttct caggagctat cagaaactat
     5641 cctcaccatg gtagccaatt gcagtaatgt tatgaattaa ttctagagct agccctagga
     5701 gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa
     5761 taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagga
     5821 ggaactatat ccggatatcc cgcaagaggc ccggcagtac cggcataacc aagcctatgc
     5881 ctacagcatc cagggtgacg gtgccgagga tgacgatgag cgcattgtta gatttcatac
     5941 acggtgcctg actgcgttag caatttaact gtgataaact accgcattaa agcttatcga
     6001 tgataagctg tcaaacatga gaattacaac ttatatcgta tggggctgac ttcaggtgct
     6061 acatttgaag agataaattg cactgaaatc aagaaatatt ttatctgatt aataagatga
     6121 tcttcttgag atcgttttgg tctgcgcgta atctcttgct ctgaaaacga aaaaaccgcc
     6181 ttgcagggcg gtttttcgaa ggttctctga gctaccaact ctttgaaccg aggtaactgg
     6241 cttggaggag cgcagtcacc aaaacttgtc ctttcagttt agccttaacc ggcgcatgac
     6301 ttcaagacta actcctctaa atcaattacc agtggctgct gccagtggtg cttttgcatg
     6361 tctttccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cggactgaac
     6421 ggggggttcg tgcatacagt ccagcttgga gcgaactgcc tacccggaac tgagtgtcag
     6481 gcgtggaatg agacaaacgc ggccataaca gcggaatgac accggtaaac cgaaaggcag
     6541 gaacaggaga gcgcacgagg gagccgccag ggggaaacgc ctggtatctt tatagtcctg
     6601 tcgggtttcg ccaccactga tttgagcgtc agatttcgtg atgcttgtca ggggggcgga
     6661 gcctatggaa aaacggcttt gccgcggccc tctcacttcc ctgttaagta tcttcctggc
     6721 atcttccagg aaatctccgc cccgttcgta agccatttcc gctcgccgca gtcgaacgac
     6781 cgagcgtagc gagtcagtga gcgaggaagc ggaatatatc ctgtatcaca tattctgctg
     6841 acgcaccggt gcagcctttt ttctcctgcc acatgaagca cttcactgac accctcatca
     6901 gtgccaacat agtaagccag tatacactcc cctagcgctg atgtccggcg gtgcttttgc
     6961 cgttacgcac caccccgtca gtagctgaac aggagggaca gctgatagaa acagaagcca
     7021 ctggagcacc tcaaaaacac catcatacac taaatcagta agttggcagc atcacccgac
     7081 gcactttgcg ccgaataaat acctgtgacg gaagatcact tcgcagaata aataaatcct
     7141 ggtgtccctg ttgataccgg gaagccctgg gccaactttt ggcgaaaatg agacgttgat
     7201 cggcacgtaa gaggttccaa ctttcaccat aatgaaataa gatcactacc gggcgtattt
     7261 tttgagttat cgagattttc aggagctaag gaagctaaaa tggagaaaaa aatcactgga
     7321 tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc atttcagtca
     7381 gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt tttaaagacc
     7441 gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc ccgcctgatg
     7501 aatgctcatc cggaattc
//
