LOCUS       sequence-199646-        7623 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The plasmid is used to express Staphylococcus aureus PcrA, a DNA 
            helicase, with N-terminal His-tag and Gly-Gly linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7623)
  TITLE     Webb Lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7623)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pET22B_SA_PcrA
FEATURES             Location/Qualifiers
     source          1..7623
                     /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             92..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2355..2372
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2425..2443)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2439..2486
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2523..2978
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2610..2629)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2820..2841
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3004..3108
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3109..3177,3178..3969)
                     /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(3327..3346)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4140..4728
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4629..4648
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4882..4899
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4914..5053
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4979..4998)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5139..5161
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5155..5346)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6155..7237)
                     /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     7199..7218
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7238..7315)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7521..7540
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcatcatcac catcaccacg gcggcatgaa
      121 tgcactgctg aatcacatga ataccgaaca gagcgaagca gttaaaacca ccgaaggtcc
      181 gctgctgatt atggcaggcg caggtagcgg taaaacccgt gttctgaccc atcgtattgc
      241 atatctgctg gatgaaaaag atgtgagccc gtataatgtt ctggcaatta cctttaccaa
      301 taaagcagcc cgtgaaatga aagaacgtgt tcagaaactg gttggtgatc aggccgaagt
      361 tatttggatg agcacctttc atagcatgtg tgttcgtatt ctgcgtcgtg atgcagatcg
      421 tattggtatt gaacgcaact ttaccattat tgatccgacc gatcagaaaa gcgttgttaa
      481 agatgtgctg aaaaacgaga acatcgatag caaaaaattc gagcctcgca tgtttattgg
      541 tgccattagc aatctgaaaa atgaactgaa aacaccggca gatgcacaga aagaagcaac
      601 cgattatcat agccagatgg ttgccaccgt ttatagcggt tatcagcgtc agctgagccg
      661 taatgaagca ctggattttg atgatctgat tatgaccacc atcaacctgt ttgaacgtgt
      721 gccggaagtt ctggaatatt atcagaacaa attccagtat atccatgtgg atgagtatca
      781 ggatacaaac aaagcacagt ataccctggt taaactgctg gcaagcaaat tcaaaaatct
      841 gtgcgttgtt ggtgatagcg atcagagcat ttatggttgg cgtggtgcag atattcagaa
      901 cattctgagc ttcgaaaaag attatccgga agccaatacc atctttctgg aacagaatta
      961 tcgtagcacc aaaaccattc tgaatgcagc caatgaagtg atcaaaaata acagcgaacg
     1021 taaaccgaaa ggtctgtgga ccgcaaatac caatggtgaa aaaatccact attatgaggc
     1081 aatgaccgaa cgtgatgaag ccgaatttgt tattcgcgaa atcatgaaac atcagcgcaa
     1141 cggcaaaaaa taccaggata tggcaattct gtatcgtacc aatgcacaga gccgtgtgct
     1201 ggaagaaacc tttatgaaaa gcaatatgcc gtataccatg gtgggtggtc agaaattcta
     1261 tgatcgcaaa gaaatcaaag acctgctgag ctatctgcgc attattgcaa atagcaacga
     1321 tgatattagc ctgcagcgta ttatcaatgt tccgaaacgt ggtgttggtc cgagcagcgt
     1381 tgaaaaagtt cagaattatg cactgcagaa caacatcagc atgtttgatg cactgggtga
     1441 agcagatttc attggtctga gcaaaaaagt tacccaagaa tgcctgaact tctatgaact
     1501 gattcagagc ctgatcaaag aacaagaatt cctggaaatc catgagatcg ttgatgaagt
     1561 gctgcagaaa agtggttatc gtgaaatgct ggaacgtgaa aataccctgg aaagccgtag
     1621 ccgtctggaa aatattgatg aatttatgag cgtgccgaaa gattatgaag aaaatacacc
     1681 tctggaagaa cagtccctga ttaactttct gaccgatctg agcctggttg cagatatcga
     1741 tgaagcggat accgaaaatg gtgttaccct gatgaccatg catagcgcaa aaggtctgga
     1801 atttccgatt gtgtttatta tgggtatgga agaaagcctg tttccgcata ttcgtgcaat
     1861 taaaagcgaa gatgatcacg agatgcaaga agaacgtcgt atttgttatg ttgcaattac
     1921 ccgtgcagaa gaggttctgt atatcaccca tgcaaccagc cgtatgctgt ttggtcgtcc
     1981 gcagagtaat atgccgagcc gttttctgaa agaaattccg gaaagcctgc tggaaaatca
     2041 tagcagcggt aaacgtcaga ccattcagcc gaaagcaaaa ccgtttgcaa aacgtggttt
     2101 tagccagcgt accaccagta ccaaaaaaca ggttctgagc agcgattgga atgtgggtga
     2161 taaagtgatg cataaagcat ggggtgaagg tatggttagc aatgtgaatg aaaaaaacgg
     2221 cagcatcgaa ctggacatca tctttaaaag ccagggtccg aaacggctgc tggcacagtt
     2281 tgcaccgatt gaaaaaaaag aggactaagg atccgaattc gagctccgtc gacaagcttg
     2341 cggccgcact cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa
     2401 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     2461 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg
     2521 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2581 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2641 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     2701 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     2761 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     2821 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     2881 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     2941 taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat
     3001 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     3061 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     3121 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     3181 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     3241 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     3301 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     3361 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     3421 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     3481 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     3541 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     3601 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg
     3661 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     3721 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     3781 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     3841 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     3901 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     3961 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     4021 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4081 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4141 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4201 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4261 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4321 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4381 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4441 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4501 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4561 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4621 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4681 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4741 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     4801 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     4861 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     4921 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta
     4981 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
     5041 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     5101 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     5161 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     5221 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     5281 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     5341 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
     5401 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
     5461 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
     5521 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
     5581 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
     5641 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
     5701 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
     5761 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc
     5821 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa
     5881 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc
     5941 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac
     6001 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca
     6061 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
     6121 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     6181 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     6241 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca
     6301 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa
     6361 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt
     6421 atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg
     6481 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca
     6541 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta
     6601 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg
     6661 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat
     6721 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct
     6781 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg
     6841 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat
     6901 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc
     6961 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca
     7021 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg
     7081 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt
     7141 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg
     7201 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct
     7261 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
     7321 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg
     7381 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
     7441 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
     7501 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
     7561 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga
     7621 aat
//
