LOCUS       sequence-233674-        6741 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows Gateway cloning of gene of interest into bacterial expression
            vector with PreScission Protease cleavable N-terminal GST tag, and a
            retained C-terminal Myc tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6741)
  TITLE     Bacterial expression plasmids for production of recombinant proteins
            with a PreScission Protease cleavable GST tag and additional small 
            affinity tags
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6741)
  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            "Unpublished"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: pGEX6P1-DEST-Myc
FEATURES             Location/Qualifiers
     source          1..6741
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..17
                     /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             40..693
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     651..673
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             700..723
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     protein_bind    748..872
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(897..902,903..920,921..927)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             981..1640
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1047..1066)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1982..2287
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2166..2185
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2328..2452)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2475..2504
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2573..2595)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2740..2758)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2826..2930
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2931..2999,3000..3791)
                     /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(3149..3168)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3962..4550
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4451..4470
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4704..4721
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4794..4871
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4872..5954
                     /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(4891..4910)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6003..6008,6009..6026,6027..6033)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6041..6057
                     /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     6065..6081
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6065..6081
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             6077..6604
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6097..6114)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6097..6113)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6106..6128)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(6706..6711,6712..6727,6728..6734)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
ORIGIN
        1 ttgtgagcgg ataacaattt cacacaggaa acagtattca tgtcccctat actaggttat
       61 tggaaaatta agggccttgt gcaacccact cgacttcttt tggaatatct tgaagaaaaa
      121 tatgaagagc atttgtatga gcgcgatgaa ggtgataaat ggcgaaacaa aaagtttgaa
      181 ttgggtttgg agtttcccaa tcttccttat tatattgatg gtgatgttaa attaacacag
      241 tctatggcca tcatacgtta tatagctgac aagcacaaca tgttgggtgg ttgtccaaaa
      301 gagcgtgcag agatttcaat gcttgaagga gcggttttgg atattagata cggtgtttcg
      361 agaattgcat atagtaaaga ctttgaaact ctcaaagttg attttcttag caagctacct
      421 gaaatgctga aaatgttcga agatcgttta tgtcataaaa catatttaaa tggtgatcat
      481 gtaacccatc ctgacttcat gttgtatgac gctcttgatg ttgttttata catggaccca
      541 atgtgcctgg atgcgttccc aaaattagtt tgttttaaaa aacgtattga agctatccca
      601 caaattgata agtacttgaa atccagcaag tatatagcat ggcctttgca gggctggcaa
      661 gccacgtttg gtggtggcga ccatcctcca aaatcggatc tggaagttct gttccagggg
      721 cccctgggat ccccggaatt cccatcaaca agtttgtaca aaaaagctga acgagaaacg
      781 taaaatgata taaatatcaa tatattaaat tagattttgc ataaaaaaca gactacataa
      841 tactgtaaaa cacaacatat ccagtcacta tggcggccgc attaggcacc ccaggcttta
      901 cactttatgc ttccggctcg tataatgtgt ggattttgag ttaggatccg tcgagatttt
      961 caggagctaa ggaagctaaa atggagaaaa aaatcactgg atataccacc gttgatatat
     1021 cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa tgtacctata
     1081 accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa aataagcaca
     1141 agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat ccggaattcc
     1201 gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg
     1261 ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc
     1321 ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt
     1381 tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg gtgagtttca
     1441 ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcaccatgg
     1501 gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg
     1561 ccgtttgtga tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg
     1621 agtggcaggg cggggcgtaa agatctggat ccggcttact aaaagccaga taacagtatg
     1681 cgtatttgcg cgctgatttt tgcggtataa gaatatatac tgatatgtat acccgaagta
     1741 tgtcaaaaag aggtatgcta tgaagcagcg tattacagtg acagttgaca gcgacagcta
     1801 tcagttgctc aaggcatata tgatgtcaat atctccggtc tggtaagcac aaccatgcag
     1861 aatgaagccc gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga agggatggct
     1921 gaggtcgccc ggtttattga aatgaacggc tcttttgctg acgagaacag gggctggtga
     1981 aatgcagttt aaggtttaca cctataaaag agagagccgt tatcgtctgt ttgtggatgt
     2041 acagagtgat attattgaca cgcccgggcg acggatggtg atccccctgg ccagtgcacg
     2101 tctgctgtca gataaagtct cccgtgaact ttacccggtg gtgcatatcg gggatgaaag
     2161 ctggcgcatg atgaccaccg atatggccag tgtgccggtc tccgttatcg gggaagaagt
     2221 ggctgatctc agccaccgcg aaaatgacat caaaaacgcc attaacctga tgttctgggg
     2281 aatataaatg tcaggctccc ttatacacag ccagtctgca ggtcgaccat agtgactgga
     2341 tatgttgtgt tttacagtat tatgtagtct gttttttatg caaaatctaa tttaatatat
     2401 tgatatttat atcattttac gtttctcgtt cagctttctt gtacaaagtg gttgatgggt
     2461 cgactcgaag cggtgaacag aaactgatca gcgaagagga tctgtaactc gagcggccgc
     2521 atcgtgactg actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac
     2581 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     2641 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac
     2701 gtagcgatag cggagtgtat aattcttgaa gacgaaaggg cctcgtgata cgcctatttt
     2761 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     2821 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     2881 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     2941 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     3001 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     3061 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     3121 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg
     3181 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     3241 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     3301 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     3361 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     3421 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa
     3481 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     3541 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     3601 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     3661 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     3721 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     3781 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     3841 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     3901 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     3961 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     4021 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     4081 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     4141 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4201 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4261 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4321 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     4381 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     4441 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     4501 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     4561 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     4621 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     4681 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     4741 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat tccgacacca
     4801 tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
     4861 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
     4921 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
     4981 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
     5041 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
     5101 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
     5161 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
     5221 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
     5281 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca
     5341 acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat
     5401 tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc
     5461 gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg
     5521 aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca
     5581 tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca
     5641 ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg
     5701 aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg
     5761 ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc
     5821 agctgttgcc cgtctcactg gtgaaaagaa aaaccacgct agcgcccaat acgcaaaccg
     5881 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     5941 aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcaccccag
     6001 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     6061 cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt tacaacgtcg
     6121 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
     6181 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     6241 gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg aaagctggct
     6301 ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc agatgcacgg
     6361 ttacgatgcg cccatctaca ccaacgtgac ctatcccatt acggtcaatc cgccgtttgt
     6421 tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg aaagctggct
     6481 acaggaaggc cagacgcgaa ttatttttga tggcgttgga attagcttat cgactgcacg
     6541 gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc
     6601 gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt
     6661 gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac aattaatcat
     6721 cggctcgtat aatgtgtgga a
//
