LOCUS       sequence-432470-        7186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PaFtsH2-6xHis cloned in XbaI/NheI site in pET-28a(+).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7186)
  AUTHORS   Mawla GD, Kamal SM, Cao LY, Purhonen P, Hebert H, Sauer RT, Baker 
            TA, Romling U
  TITLE     The membrane-cytoplasmic linker defines activity of FtsH proteases 
            in Pseudomonas aeruginosa clone C.
  JOURNAL   J Biol Chem. 2024 Feb;300(2):105622. doi: 10.1016/j.jbc.2023.105622.
            Epub 2024 Jan 3.
  PUBMED    38176647
REFERENCE   2  (bases 1 to 7186)
  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: "J Biol 
            Chem."; date: "2024-02"; volume: "300(2):105622. doi"; pages: " 
            10.1016/j.jbc.2023.105622. Epub 2024 Jan 3"
COMMENT     Sequence Label: pSG059
FEATURES             Location/Qualifiers
     source          1..7186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(205..234)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="ASMTGGQQMG"
     CDS             complement(238..255)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    2164..2188
                     /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)."
     primer_bind     complement(2188..2207)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2189..2207)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2291..2310)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2516..2593
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2594..3676
                     /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"
     CDS             4485..4676
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4670..4692)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4778..4917
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4833..4852
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4932..4949)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5103..5691)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5183..5202)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5802..5821)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5813..6628
                     /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             5813..6628
                     /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     complement(5886..5905)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(6720..7175)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6857..6878)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7069..7088
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccgacccat ttgctgtcca ccagtcatgc tagctcagtg
      241 atgatgatga tgatgtggtg tggacccttg gggtgtagag agtgaggttt cgggctggag
      301 gggctccgac gcgcgggatt cggactgcaa gccctgcgtc agttggcgga tgtcgttctc
      361 gtcgagcgtt tcccgcgcca gcagttcgcg cgcacaacgc tccagcaccg cgcggttggt
      421 gtcgaggatc cggtaggcgc gctcgaacac gcccatcacg atgtcgcgga tggcctgatc
      481 gatgcgcgcc tgggtcgatt cggccacccg gcagccgccg tgggccagtt ccggtgtatc
      541 gagaaagcgg ggccgctgcg cctcgaaggc gatgtagccc agtccttcgt ccatgccaaa
      601 gcgggtgatc atgtcgcggg caatgtcggt ggcccgtgcg aggtcatcgg cagccccggt
      661 ggacagctcg ccgaacacga gcttctcggc cgcacgaccg cccagcagca cggcgatctt
      721 gtgttccaga tcggcgcgcg tcatcaggaa gcggtcttcg gtggggcgtt gcagcgtgta
      781 gcccagcgcg cccatgccgc gcgggatgat cgagatcttg tgcacggggt cggtgccggg
      841 cagcgccagc gccaccagcg catggcccat ctcgtgatag gccacggttt cccgctcctt
      901 gggattgagc actcggttct tcttctccag gcccgccacg atgcgctcga tggcggcggt
      961 gaaatcctgc aactccacgg cggacgcttt gcgccgggtc gcggccagcg cggcctcgtt
     1021 gaccaggttc gcgaggtctg cacccgaaaa gcctgtggtc agcgcagcca cctgttcgag
     1081 atcgatatcc tgagccagcg tcaccttctt gacgtggact ttcaggatgt ccagccgtcc
     1141 cttcttgtcg ggccggtcca ccagcacctg gcggtcgaag cggccggcac gcagcagcgc
     1201 ctggtcgagg atttcggggc ggttggtggc ggcgaggatg atcagcccca ccgagctgtc
     1261 gaagccgtcc atctcggtga gcagctggtt gagcgtctgc tcacgctcgt cgtggccgcc
     1321 gatgggcccg ccgacgccgc gtgcacggcc cagcgcgtcg agttcatcga tgaagatgat
     1381 ggccggcgcc tgcccgcgcg cctgctcgaa caggtcgcgc acccgcgccg cgcccacgcc
     1441 gacaaacatc tcgacgaact ccgagccgga gatagagaag aacggtaccc cggcctcgcc
     1501 cgccacggcc ttggccagca gggtcttgcc cgtgccgggc gggccaacca gcaacacacc
     1561 tttcgggatg cgcgcgccga ggcggccgta atcctgcggg ttcttcagga agtcgacgat
     1621 ttcgaccagc tcggccttgg cctcatcgac acccgcgaca tcggcaaagg tgacgccggt
     1681 gttcttctcc atgaacacct tggcgcggct cttgccgatg ctcaggaagc cgcccatgcc
     1741 ctgcttctcg gcgaagcggc ggaacaggaa gaaccagacg ccgaagaagg ccaccgccgg
     1801 cagaatccag gagagcacat cacgtagcca ggtgctttcc accacccgcg cgtaaggcac
     1861 gtcgtacttg gacagccgct cggccaggtc gggttcgacg cgagtggcca cgatggtggt
     1921 cttgccccgg ctgtccggcg atttcaggcg cccggtgacc gtgcggtccg acaccagcac
     1981 ttcggcgacg cgcccctcgg ccagcgcctt ctcgaattcg ctgtagggca cgggctcgac
     2041 ggtcttggcc gcctgccagt agttctgcag cgtcagcagc aacaggccgg cgacgatcca
     2101 gtagccaatg ttccattgat ctttcttttc catgggcaat gttcctcgca agtcgtctag
     2161 aggggaattg ttatccgctc acaattcccc tatagtgagt cgtattaatt tcgcgggatc
     2221 gagatctcga tcctctacgc cggacgcatc gtggccggca tcaccggcgc cacaggtgcg
     2281 gttgctggcg cctatatcgc cgacatcacc gatggggaag atcgggctcg ccacttcggg
     2341 ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc ccgtggccgg gggactgttg
     2401 ggcgccatct ccttgcatgc accattcctt gcggcggcgg tgctcaacgg cctcaaccta
     2461 ctactgggct gcttcctaat gcaggagtcg cataagggag agcgtcgaga tcccggacac
     2521 catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt
     2581 cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc
     2641 ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga
     2701 aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact
     2761 ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc
     2821 gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt
     2881 gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc
     2941 gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt
     3001 ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat
     3061 caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc atctggtcgc
     3121 attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct
     3181 gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg
     3241 ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg
     3301 catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc
     3361 cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac
     3421 cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct
     3481 ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa
     3541 tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac
     3601 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
     3661 ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat taggcaccgg
     3721 gatctcgacc gatgcccttg agagccttca acccagtcag ctccttccgg tgggcgcggg
     3781 gcatgactat cgtcgccgca cttatgactg tcttctttat catgcaactc gtaggacagg
     3841 tgccggcagc gctctgggtc attttcggcg aggaccgctt tcgctggagc gcgacgatga
     3901 tcggcctgtc gcttgcggta ttcggaatct tgcacgccct cgctcaagcc ttcgtcactg
     3961 gtcccgccac caaacgtttc ggcgagaagc aggccattat cgccggcatg gcggccccac
     4021 gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg ggttgcctta
     4081 ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc tgctgcaaaa
     4141 cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg taaagtctgg
     4201 aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca ggatgctgct
     4261 ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga ccctgagtga
     4321 tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa cgttccagta
     4381 accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt ttcatcggta
     4441 tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc aaacaggaaa
     4501 aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt ctggagaaac
     4561 tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac gaccacgctg
     4621 atgagcttta ccgcagctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
     4681 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
     4741 gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta
     4801 gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg tactgagagt
     4861 gcaccattgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
     4921 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4981 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     5041 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     5101 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     5161 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     5221 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     5281 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     5341 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     5401 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     5461 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5521 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
     5581 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5641 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5701 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5761 gtcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     5821 tattcaacgg gaaacgtctt gctctaggcc gcgattaaat tccaacatgg atgctgattt
     5881 atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt
     5941 gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa
     6001 tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac
     6061 catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg
     6121 gaaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc
     6181 gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag
     6241 cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc
     6301 gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca
     6361 taaacttttg ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa
     6421 ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc
     6481 agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt
     6541 acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt
     6601 tcatttgatg ctcgatgagt ttttctaaga attaattcat gagcggatac atatttgaat
     6661 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaccta
     6721 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     6781 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     6841 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     6901 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     6961 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     7021 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     7081 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     7141 acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat tcgcca
//
