LOCUS       sequence-233662-        5020 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Allows cloning of gene of interest into bacterial expression vector 
            with PreScission Protease cleavable N-terminal GST tag, and a 
            retained N-terminal 10x His tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5020)
  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  JOURNAL   Unpublished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 5020)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pGEX6P1-N-His
FEATURES             Location/Qualifiers
     source          1..5020
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    8..24
                     /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             47..700
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     658..680
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             707..730
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             740..769
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(859..881)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1026..1044)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1112..1216
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1217..1285,1286..2077)
                     /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(1435..1454)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2248..2836
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2737..2756
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2990..3007
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3080..3157
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3158..4240
                     /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(3177..3196)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4289..4294,4295..4312,4313..4319)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4327..4343
                     /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     4351..4367
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4351..4367
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4363..4890
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4383..4400)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4383..4399)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4392..4414)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(4992..4997,4998..5013,5014..5020)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
ORIGIN
        1 tgtggaattg tgagcggata acaatttcac acaggaaaca gtattcatgt cccctatact
       61 aggttattgg aaaattaagg gccttgtgca acccactcga cttcttttgg aatatcttga
      121 agaaaaatat gaagagcatt tgtatgagcg cgatgaaggt gataaatggc gaaacaaaaa
      181 gtttgaattg ggtttggagt ttcccaatct tccttattat attgatggtg atgttaaatt
      241 aacacagtct atggccatca tacgttatat agctgacaag cacaacatgt tgggtggttg
      301 tccaaaagag cgtgcagaga tttcaatgct tgaaggagcg gttttggata ttagatacgg
      361 tgtttcgaga attgcatata gtaaagactt tgaaactctc aaagttgatt ttcttagcaa
      421 gctacctgaa atgctgaaaa tgttcgaaga tcgtttatgt cataaaacat atttaaatgg
      481 tgatcatgta acccatcctg acttcatgtt gtatgacgct cttgatgttg ttttatacat
      541 ggacccaatg tgcctggatg cgttcccaaa attagtttgt tttaaaaaac gtattgaagc
      601 tatcccacaa attgataagt acttgaaatc cagcaagtat atagcatggc ctttgcaggg
      661 ctggcaagcc acgtttggtg gtggcgacca tcctccaaaa tcggatctgg aagttctgtt
      721 ccaggggccc ctgggatccc atcatcatca tcatcatcat catcatcacc gatccccgga
      781 attcccgggt cgactcgagc ggccgcatcg tgactgactg acgatctgcc tcgcgcgttt
      841 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct
      901 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg
      961 tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtataatt cttgaagacg
     1021 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     1081 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     1141 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     1201 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     1261 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     1321 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     1381 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     1441 tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     1501 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     1561 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     1621 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     1681 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     1741 gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga
     1801 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     1861 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     1921 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     1981 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     2041 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     2101 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     2161 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     2221 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     2281 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     2341 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     2401 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     2461 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     2521 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     2581 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     2641 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     2701 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     2761 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     2821 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     2881 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     2941 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3001 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     3061 ttcacaccgc ataaattccg acaccatcga atggcgcaaa acctttcgcg gtatggcatg
     3121 atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga
     3181 tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag
     3241 ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat
     3301 tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac
     3361 ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga
     3421 tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa
     3481 agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct
     3541 ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct
     3601 tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg
     3661 actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc
     3721 attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa
     3781 tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca
     3841 aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca
     3901 gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat
     3961 ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac
     4021 catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc
     4081 tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac
     4141 cacgctagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     4201 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtaa
     4261 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     4321 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacggat
     4381 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     4441 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     4501 cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gctttgcctg gtttccggca
     4561 ccagaagcgg tgccggaaag ctggctggag tgcgatcttc ctgaggccga tactgtcgtc
     4621 gtcccctcaa actggcagat gcacggttac gatgcgccca tctacaccaa cgtgacctat
     4681 cccattacgg tcaatccgcc gtttgttccc acggagaatc cgacgggttg ttactcgctc
     4741 acatttaatg ttgatgaaag ctggctacag gaaggccaga cgcgaattat ttttgatggc
     4801 gttggaatta gcttatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg
     4861 gaagctgtgg tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg
     4921 cactcccgtt ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct
     4981 gaaatgagct gttgacaatt aatcatcggc tcgtataatg
//
