LOCUS       sequence-349047-        6035 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression plasmid for mutant R4-4 of Sfp phosphopantetheinyl 
            transferase. R4-4 can use 3′-dephospho CoA for adding the 
            phosphopantetheinyl group to the carrier proteins..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6035)
  AUTHORS   Sunbul M, Marshall NJ, Zou Y, Zhang K, Yin J
  TITLE     Catalytic turnover-based phage selection for engineering the 
            substrate specificity of Sfp phosphopantetheinyl transferase.
  JOURNAL   J Mol Biol. 2009 Apr 10;387(4):883-98. doi: 
            10.1016/j.jmb.2009.02.010.
  PUBMED    19340948
REFERENCE   2  (bases 1 to 6035)
  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.jmb.2009.02"; journalName: "J Mol Biol"; date: 
            "2009-04-10- 10"; volume: "387"; issue: "4"; pages: "883-98"
COMMENT     Sequence Label: pET-Sfp R4-4 mutant
FEATURES             Location/Qualifiers
     source          1..6035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     965..984
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1004..1081)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1287..1306
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1390..1409
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1390..1408
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1409..1433
                     /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             1448..1470
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2156..2173
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2226..2244)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2240..2287
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2324..2779
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2411..2430)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2621..2642
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2805..2909
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2910..2978,2979..3770)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3128..3147)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3941..4529
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4430..4449
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4683..4700
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4715..4854
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4780..4799)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4940..4962
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4956..5147)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5956..6035,1..1003))
                     /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"
ORIGIN
        1 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
       61 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
      121 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
      181 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
      241 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
      301 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
      361 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
      421 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
      481 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
      541 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
      601 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
      661 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
      721 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
      781 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
      841 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
      901 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
      961 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     1021 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     1081 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     1141 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     1201 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     1261 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     1321 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
     1381 ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag
     1441 aaataatttt gtttaacttt aagaaggaga tatacatatg aagatttacg gaatttatat
     1501 ggaccgcccg ctttcacagg aagaaaatga acggttcatg actttcatat cacctgaaga
     1561 gcgggagaag tgtcgacgat tttatcataa agaagatgct caccgcgagc tcctaggaga
     1621 tgtgctcgtt cgctcagtca taagcaggca gtatcagttg gacaaatccg atatccgctt
     1681 tagcacgcag gaatacggga agccgtacat ccctgatctt cccgacgctc atttcaacat
     1741 ttctcattcc ggccgctggg tcattggtgc gtttgattca cagccgatcg gcatagatat
     1801 cgaaaaaacg aaaccgatca gccttgagat cgccaagcgc ttcttttcaa aaacagagta
     1861 cagcgacctt ttagcaaaag acaaggacga gcagacagac tatttttatc atctatggtc
     1921 aatgaaagaa agctttatca aacaggaagg caaaggctta tcgcttccgc ttgattcctt
     1981 ttcagtgcgc ctgcatcagg acggacaagt atccattgag cttccggaca gccattcccc
     2041 atgctatatc aaaacgtatg aggtcgatcc cggctacaaa atggctgtat gcgccgcaca
     2101 ccctgatttc cccgaggata tcacaatggt ctcgtacgaa gagcttttac tcgagcacca
     2161 ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc
     2221 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
     2281 ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc
     2341 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2401 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     2461 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     2521 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     2581 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     2641 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     2701 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     2761 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     2821 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2881 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2941 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     3001 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     3061 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     3121 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     3181 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     3241 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     3301 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     3361 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     3421 taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac
     3481 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     3541 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     3601 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     3661 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     3721 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     3781 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     3841 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     3901 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     3961 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4021 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4081 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4141 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4201 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4261 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4321 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4381 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4441 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4501 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4561 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4621 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     4681 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     4741 atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc
     4801 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
     4861 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     4921 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     4981 aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg
     5041 tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt
     5101 ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg
     5161 taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg
     5221 atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg
     5281 gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt
     5341 aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac
     5401 ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag
     5461 accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc
     5521 tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc
     5581 ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc gataatggcc
     5641 tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc
     5701 aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc
     5761 tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag
     5821 acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg
     5881 ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca
     5941 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     6001 taatgaatcg gccaacgcgc ggggagaggc ggttt
//
