LOCUS       sequence-377729-        6918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminal GST-(His)6-Thrombin cleavage site fusion of 
            human gene or portion of gene in bacterial strains. pET based 
            vector..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6918)
  TITLE     Davis Lab plasmids
REFERENCE   2  (bases 1 to 6918)
  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: PRPF4.211.522.gst
FEATURES             Location/Qualifiers
     source          1..6918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             89..742
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             770..787
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             797..814
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1783..1800
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1853..1871)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1867..1914
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1951..2406
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2038..2057)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2248..2269
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2498..3313)
                     /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(2498..3313)
                     /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     3221..3240
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3305..3324
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3435..4023
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3924..3943
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4177..4194
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4209..4348
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4274..4293)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4434..4456
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4450..4641)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5450..6532)
                     /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     6494..6513
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6533..6610)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6816..6835
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gtcccctata ctaggttatt ggaaaattaa
      121 gggccttgtg caacccactc gacttctttt ggaatatctt gaagaaaaat atgaagagca
      181 tttgtatgag cgcgatgaag gtgataaatg gcgaaacaaa aagtttgaat tgggtttgga
      241 gtttcccaat cttccttatt atattgatgg tgatgttaaa ttaacacagt ctatggccat
      301 catacgttat atagctgaca agcacaacat gttgggtggt tgtccaaaag agcgtgcaga
      361 gatttcaatg cttgaaggag cggttttgga tattagatac ggtgtttcga gaattgcata
      421 tagtaaagac tttgaaactc tcaaagttga ttttcttagc aagctacctg aaatgctgaa
      481 aatgttcgaa gatcgtttat gtcataaaac atatttaaat ggtgatcatg taacccatcc
      541 tgacttcatg ttgtatgacg ctcttgatgt tgttttatac atggacccaa tgtgcctgga
      601 tgcgttccca aaattagttt gttttaaaaa acgtattgaa gctatcccac aaattgataa
      661 gtacttgaaa tccagcaagt atatagcatg gcctttgcag ggctggcaag ccacgtttgg
      721 tggtggcgac catcctccaa aatcggatgg ttcatccatg ggcagcagcc atcatcatca
      781 tcatcacagc agcggcctgg ttccgcgtgg tagtatgcaa gagctgcaca agtctctccg
      841 gtctttgaat aatttttgca gtcagattgg ggatgatcgg cctatctcct actgtcactt
      901 tagtcccaat tccaagatgc tggccacagc ttgttggagt gggctttgca agctctggtc
      961 tgttcctgat tgcaacctcc ttcacactct tcgagggcat aacacaaatg taggagcaat
     1021 tgtattccat cccaaatcca ctgtctcctt ggacccaaaa gatgtcaacc tggcctcttg
     1081 tgcggctgat ggctctgtga agctttggag tctcgacagt gatgaaccag tggcagatat
     1141 tgaaggccat acagtgcgtg tggcgcgggt aatgtggcat ccttcaggac gtttcctggg
     1201 caccacctgc tatgaccgtt catggcgctt atgggatttg gaggctcaag aggagatcct
     1261 gcatcaggaa ggccatagca tgggtgtgta tgacattgcc ttccatcaag atggctcttt
     1321 ggctggcact gggggactgg atgcatttgg tcgagtttgg gacctacgca caggacgttg
     1381 tatcatgttc ttagaaggcc acctgaaaga aatctatgga ataaatttct cccccaatgg
     1441 ctatcacatt gcaaccggca gtggtgacaa cacctgcaaa gtgtgggacc tccgacagcg
     1501 gcgttgcgtc tacaccatcc ctgctcatca gaacttagtg actggtgtca agtttgagcc
     1561 tatccatggg aacttcttgc ttactggtgc ctatgataac acagccaaga tctggacgca
     1621 cccaggctgg tccccgctga agactctggc tggccacgaa ggcaaagtga tgggcctaga
     1681 tatttcttcc gatgggcagc tcatagccac ttgctcatat gacaggacct tcaagctgtg
     1741 gatggctgaa tagtgatgac gaagcttgcg gccgcactcg agcaccacca ccaccaccac
     1801 tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     1861 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
     1921 ggaggaacta tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     1981 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2041 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2101 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2161 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2221 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2281 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2341 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     2401 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     2461 aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca tcgagcatca
     2521 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     2581 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     2641 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     2701 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     2761 gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     2821 cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc
     2881 gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     2941 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     3001 ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     3061 tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa
     3121 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     3181 catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     3241 catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac gtttcccgtt
     3301 gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc
     3361 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     3421 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     3481 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     3541 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     3601 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     3661 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     3721 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     3781 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     3841 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     3901 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     3961 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4021 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4081 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     4141 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     4201 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat
     4261 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct
     4321 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc
     4381 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     4441 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag
     4501 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc
     4561 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc
     4621 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt
     4681 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc
     4741 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag
     4801 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg
     4861 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg
     4921 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc
     4981 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
     5041 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
     5101 cacccgtggg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc
     5161 gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag
     5221 gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc
     5281 cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt
     5341 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
     5401 agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
     5461 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     5521 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
     5581 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt
     5641 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt
     5701 cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg
     5761 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa
     5821 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc
     5881 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca
     5941 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac
     6001 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac
     6061 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
     6121 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
     6181 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
     6241 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
     6301 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
     6361 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
     6421 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
     6481 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
     6541 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
     6601 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
     6661 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
     6721 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
     6781 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     6841 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
     6901 atctcgatcc cgcgaaat
//
