LOCUS       sequence-82999-e        5902 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-tagged E.coli replication terminator protein Tus..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5902)
  AUTHORS   Tome JM, Ozer A, Pagano JM, Gheba D, Schroth GP, Lis JT
  TITLE     Comprehensive analysis of RNA-protein interactions by 
            high-throughput sequencing-RNA affinity profiling.
  JOURNAL   Nat Methods. 2014 Jun;11(6):683-8. doi: 10.1038/nmeth.2970. Epub 
            2014 May 8.
  PUBMED    24809628
REFERENCE   2  (bases 1 to 5902)
  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: 
            "10.1038/nmeth.2970"; journalName: "Nat Methods"; date: "2014-06"; 
            volume: "11"; issue: "6"; pages: "683-8"
COMMENT     Sequence Label: Tus pGST-//
FEATURES             Location/Qualifiers
     source          1..5902
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             924..944
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(1952..1974)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2119..2137)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2205..2309
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2310..2378,2379..3170)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2528..2547)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3341..3929
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3830..3849
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4083..4100
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4173..4250
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4251..5333
                     /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     complement(4270..4289)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5346..5367
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5382..5387,5388..5405,5406..5412)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5420..5436
                     /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     5444..5460
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5444..5460
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5456..5902,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5476..5493)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5476..5492)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5485..5507)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa aatcgatacg accgaaaacc tgtattttca gggcgccatg gatccgatgg
      961 cgcgttacga tctcgtagac cgactcaaca ctacctttcg ccagatggaa caagagctgg
     1021 ctatatttgc cgctcatctt gagcaacaca agctattggt tgcccgcgtg ttctctttgc
     1081 cggaggtaaa aaaagaggat gagcataatc cgcttaatcg tattgaggta aaacaacatc
     1141 tcggcaacga cgcgcagtcg ctggcgttgc gtcatttccg ccatttattt attcaacaac
     1201 agtccgaaaa tcgcagcagc aaggccgctg tccgtctgcc tggcgtgttg tgttaccagg
     1261 tcgataacct ttcgcaagca gcgttggtca gtcatattca gcacatcaat aaactcaaga
     1321 ccacgttcga gcatatcgtc acggttgaat cagaactccc caccgcggca cgttttgaat
     1381 gggtgcatcg tcatttgccg gggctgatca cccttaatgc ttaccgcacg ctcaccgttc
     1441 tgcacgaccc cgccacttta cgctttggtt gggctaataa acatatcatt aagaatttac
     1501 atcgtgatga agtcctggca cagctggaaa aaagcctgaa atcaccacgc agtgtcgcac
     1561 cgtggacgcg cgaggagtgg caaagaaaac tggagcgaga gtatcaggat atcgctgccc
     1621 tgccacagaa cgcgaagtta aaaatcaaac gtccggtgaa ggtgcagccg attgcccgcg
     1681 tctggtacaa aggagatcaa aaacaagtcc aacacgcctg ccctacacca ctgattgcac
     1741 tgattaatcg ggataatggc gcgggcgtgc cggacgttgg tgagttgtta aattacgatg
     1801 ccgacaatgt gcagcaccgt tataaacctc aggcgcagcc gcttcgtttg atcattccac
     1861 ggctgcacct gtatgttgca gattaactcg agcggccgca tcgtgactga ctgacgatct
     1921 gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     1981 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     2041 gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata
     2101 attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     2161 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     2221 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2281 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2341 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     2401 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     2461 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     2521 aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag caactcggtc
     2581 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     2641 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     2701 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     2761 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     2821 taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac
     2881 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     2941 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     3001 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     3061 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     3121 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     3181 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     3241 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     3301 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     3361 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     3421 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     3481 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     3541 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     3601 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3661 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3721 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3781 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3841 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     3901 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     3961 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4021 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     4081 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     4141 atctgtgcgg tatttcacac cgcataaatt ccgacaccat cgaatggtgc aaaacctttc
     4201 gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag
     4261 taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg
     4321 tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg
     4381 agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga
     4441 ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta
     4501 aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg
     4561 tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca
     4621 ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc
     4681 cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg
     4741 aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc
     4801 tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat
     4861 atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt
     4921 ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg
     4981 ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg
     5041 ttggtgcgga tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc
     5101 cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct
     5161 tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg
     5221 tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     5281 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
     5341 gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg
     5401 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac
     5461 catgattacg gattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt
     5521 tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga
     5581 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcgctttgc
     5641 ctggtttccg gcaccagaag cggtgccgga aagctggctg gagtgcgatc ttcctgaggc
     5701 cgatactgtc gtcgtcccct caaactggca gatgcacggt tacgatgcgc ccatctacac
     5761 caacgtaacc tatcccatta cggtcaatcc gccgtttgtt cccacggaga atccgacggg
     5821 ttgttactcg ctcacattta atgttgatga aagctggcta caggaaggcc agacgcgaat
     5881 tatttttgat ggcgttggaa tt
//
