LOCUS       sequence-200715-        6990 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows expression of the fission yeast NatB complex - 
            chloramphenicol marker .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6990)
  AUTHORS   Johnson M, Coulton AT, Geeves MA, Mulvihill DP
  TITLE     Targeted amino-terminal acetylation of recombinant proteins in E. 
            coli.
  JOURNAL   PLoS One. 2010 Dec 23;5(12):e15801. doi: 
            10.1371/journal.pone.0015801.
  PUBMED    21203426
REFERENCE   2  (bases 1 to 6990)
  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: "PLoS One.";
            date: "2010-12-23"; volume: "5(12):e15801. doi"; pages: " 
            10.1371/journal.pone.0015801"
COMMENT     Sequence Label: pNatB (pACYCduet-naa20-naa25)
FEATURES             Location/Qualifiers
     source          1..6990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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)."
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        2656..2674
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2675..2699
                     /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)."
     CDS             3346..3390
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(3428..3446)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3442..3489
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3710..4369)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4284..4303
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4370..4472)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4864..4881)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4998..5543)
                     /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(5078..5097)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5768..6850)
                     /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     6812..6831
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6851..6928)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(6974..6990,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acatgcgtcg ttctgggagt aaagaatcaa caattgttta
      181 ttcagcgtta tccttggctc aggcaggacg tggtcccgag gctctagcac ttttagaacc
      241 gttaaaatcg actcccatca acagcttgga acttttagac attatacaag ctgtttatga
      301 tgatcaaaag aagggagaag aatcgtttgt attttgggaa aagttcttgc aaacttatgg
      361 aaagcaggaa aaaaacttat tagcttactt caaagcatct attcgcatca aatcattgtc
      421 tcaccaaaga aaggctgctg tagaattgca aaagaacttt ccttctagga aacatacctt
      481 atgggttatc tcatctttgt atcttttgtc caaaaagtcg gaaaatgagg ttgaacaacg
      541 tttgctgaaa gcactagctg aaaaaactgc taaactcatt ttcgaaaaac caacaggtta
      601 tatcgattct tgtgaggagt ttcatcttta tttagacgtg ttgcttttag tgggagacaa
      661 ggacagagcg ttagatgctt taatacatca agatgctgat cgctttgttg atgctgacgc
      721 cgaccttctt ttgaggaagc ttgaattgct tgcttcctgt gcgcgttggg attctttatt
      781 cacgttttcg ctttctcttt tccaaactgg gaatactgat tggaaggttt gcaaagcttt
      841 gctggatagt gcttccaatg atgattcaaa attagtccca ttaaaggatt gtatattgaa
      901 agcgttgtca accagcagca caaaaagaaa tcttcattta ctttggattg aagccagcgc
      961 acggtttttt cctgaagaac acgaatcagc attacttggc tatatcaaaa agctgtatat
     1021 gaaaccaatt gtttttgaag atttgagacc ttatctattg aagctgaatg ttgatgcaca
     1081 acaccggtta ttagatgcat ttaagttggc tgatctaggt gaatccaatg aatctcaaaa
     1141 agtcgataaa ttatatgctg aggttttgct tctgaagatc cattttttgc tctttgagtc
     1201 gtttactgca gaatcggttg tcgattacgt gcgtagatgc ttcgtcgcat ttgaaaaggg
     1261 tttgtcttta agtaaaggct tattacccac tgactttacg cacggttatg aagctttgtt
     1321 attagcagtt cattcattga tatatatgtg ggaggggaat aaggatttaa aacctgccga
     1381 aaaacaagct ttaatttttg acgcaatctg tcttttagaa aaaggcatta cttatagtca
     1441 acataacttt catttaaagc taccccttat tcgtttatat cttcttcttg atggaggctt
     1501 ccctgctgct gctaaagttt acgacactat gtccattaag caaattcaaa acgataccct
     1561 tgatcattat ctcttgactc gtgctactac ctattatcca tcttctgtta cttctcacta
     1621 tattaactca agtctcaaaa tatatggttc gaatgagttt gaaacaccag agatgatttc
     1681 tatggcttat gaagatggtg cttactctca aatcgaagat atgcgtaatt tccgatcccg
     1741 tttagaccac agtacctgga aaagcatttc tttggtcgag cgggctagga tacattatct
     1801 taccgctttt aaaccaccca aacagtatct tcccaaatgt tcatctccga aagataatcg
     1861 agatttaaag gtctttgcag attatggatc tgataaattg ccaacagttg aagaaagttt
     1921 aagaaattcg cctaaaccgg acacgctttg gatccatttg actgtcattg gtcattctct
     1981 cgttcaagac agcatagtaa acggcgattt tgaaaaggcc gttttaagtg ctaaggaaat
     2041 ggaagttttg tgcgaaaaca atgatttatc gaagcaatta acttcggaag aaattgttca
     2101 tatgaaatta ttgatccaac tgggattact ttcggtaaag gtcaagaacg gagactacga
     2161 aaattcctcg ttcgaaacga ttgaaaattt aattgagtca ttcgattatg agaactccac
     2221 gcctttaagt caattgacaa agtacactga aatcataaat gaccttatca cttgtctaaa
     2281 cagcttccta tatcacgtgt cagctaccaa gaaaaaagaa tttaccagac aataccaact
     2341 tcttaaaaac atcagcagta acaaattagg aagtatctct ggaattacga agcataaaaa
     2401 gaaggccgcc agaaaatatg tatcagaatt gttgtcaaac tcttggctga gcaatttatc
     2461 tgaaactcag gttccttacg atccgaagtt tgcaaaacaa gttggtgaag gaatgatcgg
     2521 ttcatacatt caaacgacgg acgcagtaag caagcttcca aaatttgtaa aattttaggt
     2581 cgacaagctt gcggccgcat aatgcttaag tcgaacagaa agtaatcgta ttgtacacgg
     2641 ccgcataatc gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc
     2701 catcttagta tattagttaa gtataagaag gagatataca tatgactgat acgcgaaaat
     2761 tcaaagctac tgatttattt tcatttaaca atataaattt agatcctcta acggagacct
     2821 tcaatatttc attttatctc agttatttga acaaatggcc tagcttgtgt gttgtacaag
     2881 agtctgacct atctgatcca acccttatgg gttatattat ggggaaatca gaaggaaccg
     2941 gtaaagagtg gcatactcat gttaccgcca ttacagtagc gccaaattcc cgtcgtttag
     3001 gccttgctag aacgatgatg gactacttag aaactgtagg aaattctgaa aacgcttttt
     3061 ttgttgacct gtttgtgcgt gccagcaatg cacttgcaat tgacttttac aaaggattgg
     3121 ggtattcggt ttatagacga gttattgggt actatagtaa tccacatgga aaggatgagg
     3181 attcatttga tatgcggaaa ccactttcgc gagatgttaa tagagaaagc attcgagaga
     3241 atggggaaaa tttcaaatgc agcccagctg atgtttcatt ttagggatct caattggata
     3301 tcggccggcc acgcgatcgc tgacgtcggt accctcgagt ctggtaaaga aaccgctgct
     3361 gcgaaatttg aacgccagca catggactcg tctactagcg cagcttaatt aacctaggct
     3421 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     3481 ggttttttgc tgaaacctca ggcatttgag aagcacacgg tcacactgct tccggtagtc
     3541 aataaaccgg taaaccagca atagacataa gcggctattt aacgaccctg ccctgaaccg
     3601 acgaccgggt cgaatttgct ttcgaatttc tgccattcat ccgcttatta tcacttattc
     3661 aggcgtagca accaggcgtt taagggcacc aataactgcc ttaaaaaaat tacgccccgc
     3721 cctgccactc atcgcagtac tgttgtaatt cattaagcat tctgccgaca tggaagccat
     3781 cacaaacggc atgatgaacc tgaatcgcca gcggcatcag caccttgtcg ccttgcgtat
     3841 aatatttgcc catagtgaaa acgggggcga agaagttgtc catattggcc acgtttaaat
     3901 caaaactggt gaaactcacc cagggattgg ctgagacgaa aaacatattc tcaataaacc
     3961 ctttagggaa ataggccagg ttttcaccgt aacacgccac atcttgcgaa tatatgtgta
     4021 gaaactgccg gaaatcgtcg tggtattcac tccagagcga tgaaaacgtt tcagtttgct
     4081 catggaaaac ggtgtaacaa gggtgaacac tatcccatat caccagctca ccgtctttca
     4141 ttgccatacg gaactccgga tgagcattca tcaggcgggc aagaatgtga ataaaggccg
     4201 gataaaactt gtgcttattt ttctttacgg tctttaaaaa ggccgtaata tccagctgaa
     4261 cggtctggtt ataggtacat tgagcaactg actgaaatgc ctcaaaatgt tctttacgat
     4321 gccattggga tatatcaacg gtggtatatc cagtgatttt tttctccatt ttagcttcct
     4381 tagctcctga aaatctcgat aactcaaaaa atacgcccgg tagtgatctt atttcattat
     4441 ggtgaaagtt ggaacctctt acgtgccgat caacgtctca ttttcgccaa aagttggccc
     4501 agggcttccc ggtatcaaca gggacaccag gatttattta ttctgcgaag tgatcttccg
     4561 tcacaggtat ttattcggcg caaagtgcgt cgggtgatgc tgccaactta ctgatttagt
     4621 gtatgatggt gtttttgagg tgctccagtg gcttctgttt ctatcagctg tccctcctgt
     4681 tcagctactg acggggtggt gcgtaacggc aaaagcaccg ccggacatca gcgctagcgg
     4741 agtgtatact ggcttactat gttggcactg atgagggtgt cagtgaagtg cttcatgtgg
     4801 caggagaaaa aaggctgcac cggtgcgtca gcagaatatg tgatacagga tatattccgc
     4861 ttcctcgctc actgactcgc tacgctcggt cgttcgactg cggcgagcgg aaatggctta
     4921 cgaacggggc ggagatttcc tggaagatgc caggaagata cttaacaggg aagtgagagg
     4981 gccgcggcaa agccgttttt ccataggctc cgcccccctg acaagcatca cgaaatctga
     5041 cgctcaaatc agtggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     5101 ggcggctccc tcgtgcgctc tcctgttcct gcctttcggt ttaccggtgt cattccgctg
     5161 ttatggccgc gtttgtctca ttccacgcct gacactcagt tccgggtagg cagttcgctc
     5221 caagctggac tgtatgcacg aaccccccgt tcagtccgac cgctgcgcct tatccggtaa
     5281 ctatcgtctt gagtccaacc cggaaagaca tgcaaaagca ccactggcag cagccactgg
     5341 taattgattt agaggagtta gtcttgaagt catgcgccgg ttaaggctaa actgaaagga
     5401 caagttttgg tgactgcgct cctccaagcc agttacctcg gttcaaagag ttggtagctc
     5461 agagaacctt cgaaaaaccg ccctgcaagg cggttttttc gttttcagag caagagatta
     5521 cgcgcagacc aaaacgatct caagaagatc atcttattaa tcagataaaa tatttctaga
     5581 tttcagtgca atttatctct tcaaatgtag cacctgaagt cagccccata cgatataagt
     5641 tgtaattctc atgttagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     5701 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     5761 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5821 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     5881 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     5941 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     6001 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagatg tccgcaccaa
     6061 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     6121 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     6181 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     6241 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     6301 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     6361 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     6421 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     6481 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     6541 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     6601 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     6661 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     6721 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     6781 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     6841 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     6901 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     6961 tgcattagga aattaatacg actcactata
//
