LOCUS       sequence-197509-        5906 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the PPTase Sfp from B. subtilis; loads CoA analogues onto 
            ACP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5906)
  AUTHORS   Worthington AS, Burkart MD
  TITLE     One-pot chemo-enzymatic synthesis of reporter-modified proteins.
  JOURNAL   Org Biomol Chem. 2006 Jan 7;4(1):44-6. Epub 2005 Nov 24.
  PUBMED    16357994
REFERENCE   2  (bases 1 to 5906)
  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: "Org Biomol 
            Chem."; date: "2006-01-7"; volume: "4(1)"; pages: "44-6. Epub 2005 
            Nov 24"
COMMENT     Sequence Label: Sfp pet29b C-terminal His Tag
FEATURES             Location/Qualifiers
     source          1..5906
                     /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             767..784
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(837..855)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      851..898
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      935..1390
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1022..1041)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1232..1253
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1482..2297)
                     /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(1482..2297)
                     /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     2205..2224
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2289..2308
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2419..3007
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2908..2927
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3161..3178
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3193..3332
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3258..3277)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3418..3440
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3434..3625)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4434..5516)
                     /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     5478..5497
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5517..5594)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5800..5819
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gaagatttac ggaatttata tggaccgccc
      121 gctttcacag gaagaaaatg aacggttcat gactttcata tcacctgaaa aacgggagaa
      181 atgccggaga ttttatcata aagaagatgc tcaccgcacc ctgctgggag atgtgctcgt
      241 tcgctcagtc ataagcaggc agtatcagtt ggacaaatcc gatatccgct ttagcacgca
      301 ggaatacggg aagccgtgca tccctgatct tcccgacgct catttcaaca tttctcactc
      361 cggccgctgg gtcattggtg cgtttgattc acagccgatc ggcatagata tcgaaaaaac
      421 gaaaccgatc agccttgaga tcgccaagcg cttcttttca aaaacagagt acagcgacct
      481 tttagcaaaa gacaaggacg agcagacaga ctatttttat catctatggt caatgaaaga
      541 aagctttatc aaacaggaag gcaaaggctt atcgcttccg cttgattcct tttcagtgcg
      601 cctgcatcag gacggacaag tatccattga gcttccggac agccattccc catgctatat
      661 caaaacgtat gaggtcgatc ccggctacaa aatggctgta tgcgccgcac accctgattt
      721 ccccgaggat atcacaatgg tctcgtacga agagctttta ctcgagcacc accaccacca
      781 ccactgagat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
      841 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
      901 gaaaggagga actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc
      961 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     1021 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     1081 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     1141 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     1201 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     1261 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     1321 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     1381 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     1441 tctaaataca ttcaaatatg tatccgctca tgaattaatt cttagaaaaa ctcatcgagc
     1501 atcaaatgaa actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc
     1561 cgtttctgta atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg
     1621 tatcggtctg cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca
     1681 aaaataaggt tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc
     1741 aaaagtttat gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca
     1801 aaatcactcg catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat
     1861 acgcgatcgc tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac
     1921 actgccagcg catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat
     1981 gctgttttcc cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa
     2041 tgcttgatgg tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct
     2101 gtaacatcat tggcaacgct acctttgcca tgtttcagaa acaactctgg cgcatcgggc
     2161 ttcccataca atcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta
     2221 tacccatata aatcagcatc catgttggaa tttaatcgcg gcctagagca agacgtttcc
     2281 cgttgaatat ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt
     2341 gttcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     2401 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2461 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2521 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     2581 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2641 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2701 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2761 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2821 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2881 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2941 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     3001 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     3061 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     3121 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     3181 agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3241 caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc
     3301 cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg
     3361 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     3421 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt
     3481 aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca
     3541 gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa
     3601 gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg
     3661 gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac
     3721 atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc
     3781 agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac
     3841 agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact
     3901 tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg
     3961 tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct
     4021 gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca
     4081 tgcgcacccg tggggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg
     4141 tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg
     4201 acaggccgat catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg
     4261 ctgccggcac ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga
     4321 tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg
     4381 gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc
     4441 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     4501 ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc
     4561 aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg
     4621 gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag
     4681 ctgtcttcgg tatcgtcgta tcccactacc gagatgtccg caccaacgcg cagcccggac
     4741 tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg
     4801 ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag
     4861 tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca
     4921 gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg
     4981 tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata
     5041 atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag
     5101 gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg
     5161 acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct
     5221 accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca
     5281 atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt
     5341 ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct
     5401 tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg
     5461 gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc
     5521 accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc
     5581 cattcgatgg tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca
     5641 gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga
     5701 gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc
     5761 gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc
     5821 gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat
     5881 cgagatcgat ctcgatcccg cgaaat
//
