LOCUS       sequence-315536-        5370 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of PilB with a C-terminal histag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5370)
  AUTHORS   Katrina Forest
  TITLE     Forest Lab PilB Plasmids
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 5370)
  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            "unpublished"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: pUCP20 PilB
FEATURES             Location/Qualifiers
     source          1..5370
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(9..27)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        95..199
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(200..268,269..1060)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(418..437)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1231..1819
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1720..1739
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1973..1990
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2107..2128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2143..2148,2149..2166,2167..2173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2181..2197
                     /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     2186..2208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             3924..3941
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(3941..4774)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pRO1600 from Pseudomonas aeruginosa "
                     /label=pRO1600 Rep
                     /translation="MASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     rep_origin      4788..5139
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from 
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
     primer_bind     complement(5201..5220)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5320..5342
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
       61 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
      121 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
      181 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
      241 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
      301 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
      361 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
      421 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
      481 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
      541 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
      601 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
      661 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
      721 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
      781 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
      841 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
      901 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
      961 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     1021 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     1081 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     1141 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     1201 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     1261 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     1321 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     1381 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     1441 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     1501 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     1561 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     1621 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     1681 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     1741 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     1801 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     1861 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     1921 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     1981 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     2041 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     2101 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     2161 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     2221 acgacagcat ccaactgagc ggtctgtccc gacagctcgt ccaagccaat ctactcgacg
     2281 aaaagaccgc cctgcaggcc cagacgcagg cgcagcgcaa caagctgtcg ctggtgacgc
     2341 acctggtgca gaacaagctg gtaagcggcc tggccctggc cgaattgtcc gccgagcagt
     2401 tcggcatcgc ctattgcgac ctgaacagcc tggacaggga aagctttccg cgggacgcca
     2461 tcagcgaaaa gctggtccgc cagcaccggg tcattcccct gtggcggcgg ggcaacaagt
     2521 tgttcgtagg catctccgac gccgccaacc accaggccat caacgacgtc cagttcagca
     2581 ctggcctgac gaccgaggcc attctggtcg aggacgacaa gctcggcctt gccatcgaca
     2641 agctattcga gaatgcaacc gacggcttgg caggtctcga tgacgtggac ctggaaggac
     2701 tggatgtagg cgtcaaggaa acgtccggac aagaagatac tggcgcggag gcagacgacg
     2761 ctcccgtcgt ccgcttcgtc aacaagatgc tgctggacgc tatcaaaggc gggtcttccg
     2821 acttgcactt cgagccctac gagaagatct accgggtgcg cttccgtacc gacggcatgc
     2881 ttcacgaagt ggccaagccg ccgatccagt tggccagtcg tatctctgct cgtctcaagg
     2941 taatggccgg cctggatatc tccgaacgac gcaagccgca ggacggacgg atcaagatgc
     3001 gcgtgtcgaa gaccaagtcc atcgacttcc gcgtcaacac cctgccgacc ctgtggggcg
     3061 agaagatcgt gatgcggatc ctcgactcct ccagcgcgca gatgggcatc gacgccctgg
     3121 gctacgagga ggaccagaag gaactctacc tggccgcact caagcagccg cagggcatga
     3181 tcctggtcac cggccccacc ggctcgggca agacggtatc gctgtacacc ggcctgaaca
     3241 tcctcaatac caccgacatc aacatttcca ccgccgaaga cccggtggag atcaacctgg
     3301 aaggcatcaa ccaggtcaac gtcaatccgc gccagggcat ggacttctcc caggcgctgc
     3361 gcgccttcct gcgccaggac ccggacgtga tcatggtcgg cgagatccgc gacctggaga
     3421 ccgccgagat cgccatcaag gcggcgcaga ccgggcatat ggtgatgtcc accctgcaca
     3481 ccaacagcgc cgccgagacc ctgacccgcc tgctgaacat gggcgtaccg gcgttcaacc
     3541 tggcgacctc ggtgaacctg atcatcgccc agcgccttgc gcgaaaactc tgttcgcact
     3601 gcaagaaaga gcacgacgta ccgaaggaaa ccctgcttca cgaaggtttc ccagaagagt
     3661 tgatcggcac cttcaagctg tattcaccag tgggctgcga ccattgcaag aacggttaca
     3721 agggccgtgt cggtatttat gaagtagtta aaaacacccc ggccctgcag cggattatca
     3781 tggaggaagg caactccatc gagatcgccg agcaagcccg caaagaaggc ttcaacgatc
     3841 tgcgcacctc aggcctgctg aaagccatgc aggggatcac cagcctggag gaagtcaacc
     3901 gcgtgaccaa ggataggccg ggccatcatc atcaccatca ttaggccaga tccagcggca
     3961 tctgggttag tcgagcgcgg gccgcttccc atgtctcacc agggcgagcc tgtttcgcga
     4021 tctcagcatc tgaaatcttc ccggccttgc gcttcgctgg ggccttaccc accgccttgg
     4081 cgggcttctt cggtccaaaa ctgaacaaca gatgtgtgac cttgcgcccg gtctttcgct
     4141 gcgcccactc cacctgtagc gggctgtgct cgttgatctg cgtcacggct ggatcaagca
     4201 ctcgcaactt gaagtccttg atcgagggat accggccttc cagttgaaac cactttcgca
     4261 gctggtcaat ttctatttcg cgctggccga tgctgtccca ttgcatgagc agctcgtaaa
     4321 gcctgatcgc gtgggtgctg tccatcttgg ccacgtcagc caaggcgtat ttggtgaact
     4381 gtttggtgag ttccgtcagg tacggcagca tgtctttggt gaacctgagt tctacacggc
     4441 cctcaccctc ccggtagatg attgtttgca cccagccggt aatcatcaca ctcggtcttt
     4501 tccccttgcc attgggctct tgggttaacc ggacttcccg ccgtttcagg cgcagggccg
     4561 cttctttgag ctggttgtag gaagattcga tagggacacc cgccatcgtc gctatgtcct
     4621 ccgccgtcac tgaatacatc acttcatcgg tgacaggctc gctcctcttc acctggctaa
     4681 tacaggccag aacgatccgc tgttcctgaa cactgaggcg atacgcggcc tcgaccaggg
     4741 cattgctttt gtaaaccatt gggggtgagg ccacgttcga cattccttgt gtataagggg
     4801 acactgtatc tgcgtcccac aatacaacaa atccgtccct ttacaacaac aaatccgtcc
     4861 cttcttaaca acaaatccgt cccttaatgg caacaaatcc gtcccttttt aaactctaca
     4921 ggccacggat tacgtggcct gtagacgtcc taaaaggttt aaaagggaaa aggaagaaaa
     4981 gggtggaaac gcaaaaaacg caccactacg tggccccgtt ggggccgcat ttgtgcccct
     5041 gaaggggcgg gggaggcgtc tgggcaatcc ccgttttacc agtcccctat cgccgcctga
     5101 gagggcgcag gaagcgagta atcagggtat cgaggcggat tcacccttgg cgtccaacca
     5161 gcggcaccag cggcgcctga gaggtatggt gcactctcag tacaatctgc tctgatgccg
     5221 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     5281 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     5341 ggttttcacc gtcatcaccg aaacgcgcga
//
