LOCUS       sequence-430153-        5546 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a Pseudomonas aeruginosa metallothionein fused with the 
            S-tag at the N-terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5546)
  TITLE     Shanbhag lab plasmids
REFERENCE   2  (bases 1 to 5546)
  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"
COMMENT     Sequence Label: Stag-PaMT
FEATURES             Location/Qualifiers
     source          1..5546
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     102..119
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    134..273
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(199..218)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     359..381
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(375..566)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1375..2457)
                     /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     2419..2438
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2458..2535)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2741..2760
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2848..2867
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2848..2866
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2867..2891
                     /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             2906..2928
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2939..2983
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             2999..3016
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             3254..3271
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3324..3342)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3338..3385
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3422..3877
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3509..3528)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3719..3740
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3969..4784)
                     /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(3969..4784)
                     /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     4692..4711
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4776..4795
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4906..5494
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5395..5414
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
       61 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
      121 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
      181 gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
      241 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac
      301 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
      361 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg
      421 taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc
      481 agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg ggccatgtta
      541 agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt tctgttcatg
      601 ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac tgatgatgaa
      661 catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat gcggcgggac
      721 cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt aggtgttcca
      781 cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca gggcgctgac
      841 ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag
      901 gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc
      961 tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag gagcacgatc
     1021 atgcgcaccc gtggggccgc catgccggcg ataatggcct gcttctcgcc gaaacgtttg
     1081 gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca agattccgaa taccgcaagc
     1141 gacaggccga tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat gacccagagc
     1201 gctgccggca cctgtcctac gagttgcatg ataaagaaga cagtcataag tgcggcgacg
     1261 atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc
     1321 ggtcgagatc ccggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg
     1381 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     1441 gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg
     1501 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
     1561 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga
     1621 gctgtcttcg gtatcgtcgt atcccactac cgagatgtcc gcaccaacgc gcagcccgga
     1681 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
     1741 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
     1801 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
     1861 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg
     1921 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
     1981 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
     2041 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
     2101 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
     2161 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
     2221 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
     2281 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
     2341 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
     2401 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
     2461 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg
     2521 ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc
     2581 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg
     2641 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
     2701 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
     2761 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
     2821 tcgagatcga tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg
     2881 ataacaattc ccctctagaa ataattttgt ttaactttaa gaaggagata tacatatgaa
     2941 agaaaccgct gctgctaaat tcgaacgcca gcacatggac agcccagatc tgggtaccct
     3001 ggtgccacgc ggttccatgg gcatgaacga ccagcgttgc gcgtgcaccc actgcagctg
     3061 caccgtggac gcgaacgcgc tgcaacgtga tggcaaggcg tactgctgcg aggcgtgcgc
     3121 gagcggtcac cgtaaaggcg aaccgtgccg tatgcaggat tgccactgcg gcgagaagcc
     3181 gggcgaaagc gcggttgaca acgcgctgga tgagaccttc ccggcgagcg aaccgatcag
     3241 ctaataactc gagcaccacc accaccacca ctgagatccg gctgctaaca aagcccgaaa
     3301 ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc
     3361 taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat tggcgaatgg
     3421 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3481 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3541 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3601 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3661 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3721 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     3781 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3841 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     3901 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     3961 attaattctt agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga
     4021 ttatcaatac catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg
     4081 cagttccata ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca
     4141 atacaaccta ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga
     4201 gtgacgactg aatccggtga gaatggcaaa agtttatgca tttctttcca gacttgttca
     4261 acaggccagc cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt
     4321 cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt taaaaggaca attacaaaca
     4381 ggaatcgaat gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa
     4441 tcaggatatt cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac
     4501 catgcatcat caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc
     4561 agccagttta gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt
     4621 ttcagaaaca actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat
     4681 tgcccgacat tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt
     4741 aatcgcggcc tagagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta
     4801 ctgtttatgt aagcagacag ttttattgtt catgaccaaa atcccttaac gtgagttttc
     4861 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4921 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4981 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5041 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     5101 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     5161 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     5221 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     5281 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5341 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5401 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     5461 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     5521 gttcctggcc ttttgctggc cttttg
//
