LOCUS       sequence-288549-        5197 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsRck_296-472-HsPatL1_1-59.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5197)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 5197)
  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: pRSFDuetG-HsRCK_296-472-HsPatL1_1-59 (MCSII)_D
FEATURES             Location/Qualifiers
     source          1..5197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             110..133
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     promoter        742..760
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    761..785
                     /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             831..1484
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1442..1464
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1491..1514
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1734..1778
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1816..1834)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1830..1877
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2107..2922)
                     /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(2107..2922)
                     /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     2830..2849
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2923..3014)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3030..3779)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     complement(3040..3057)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3280..3299)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3975..5057)
                     /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     5019..5038
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5058..5135)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(5181..5197,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agcgcgggtc tggaagttct
      121 gttccagggg ccccaggatc cgaagggagt aacccagtac tacgcatatg taactgagcg
      181 ccaaaaagta cactgcctca acacactttt ctccaggctt cagataaacc agtcgatcat
      241 tttctgtaac tcctctcagc gagttgaatt gctagccaag aagatttctc aactgggtta
      301 ttcttgcttc tatattcatg ctaaaatgag gcaggaacat cgaaatcgtg tatttcatga
      361 tttccgaaat ggcttatgcc gcaatcttgt ttgcactgat ctgtttaccc gaggtattga
      421 tatacaagct gtgaatgtgg taataaactt tgatttccca aagctggcag agacctatct
      481 ccatcgtatt ggaagatcag gtcgctttgg tcatcttggc ttagccatca acttgatcac
      541 atatgatgat cgcttcaacc tgaaaagtat tgaggagcag ctgggaacag aaattaaacc
      601 tattccgagc aacattgata agagcctgta tgtggcagaa taccacagcg agcctgtaga
      661 agatgagaaa ccttaagcgg ccgcataatg cttaagtcga acagaaagta atcgtattgt
      721 acacggccgc ataatcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca
      781 attccccatc ttagtatatt agttaagtat aagaaggaga tatacataat atgtccccta
      841 tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc
      901 ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca
      961 aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta
     1021 aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg
     1081 gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat
     1141 acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta
     1201 gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa
     1261 atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat
     1321 acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg
     1381 aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc
     1441 agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggaagttc
     1501 tgttccaggg gccccatatg gcagatctca attggatgtt ccgctacgag tctttggagg
     1561 attgtcctct ggatgaagat gaagatgcat ttcagggact gggagaagaa gatgaagaga
     1621 ttgatcaatt caatgatgat acatttgggt caggtgcagt tgatgatgat tggcaggaag
     1681 cacatgagcg cctggctgaa ttggaagaaa agtagggtac cctcgagtct ggtaaagaaa
     1741 ccgctgctgc gaaatttgaa cgccagcaca tggactcgtc tactagcgca gcttaattaa
     1801 cctaggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     1861 tcttgagggg ttttttgctg aaacctcagg catttgagaa gcacacggtc acactgcttc
     1921 cggtagtcaa taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc
     1981 ctgaaccgac gacaagctga cgaccgggtc tccgcaagtg gcacttttcg gggaaatgtg
     2041 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat
     2101 taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt
     2161 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
     2221 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
     2281 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
     2341 gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac
     2401 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
     2461 tgattgcgcc tgagcgagac gaaatacgcg gtcgctgtta aaaggacaat tacaaacagg
     2521 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
     2581 aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca
     2641 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
     2701 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
     2761 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
     2821 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
     2881 tcgcggccta gagcaagacg tttcccgttg aatatggctc atactcttcc tttttcaata
     2941 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     3001 gaaaaataaa caaataggca tgcagcgctc ttccgcttcc tcgctcactg actcgctacg
     3061 ctcggtcgtt cgactgcggc gagcggtgtc agctcactca aaagcggtaa tacggttatc
     3121 cacagaatca ggggataaag ccggaaagaa catgtgagca aaaagcaaag caccggaaga
     3181 agccaacgcc gcaggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     3241 gacgctcaag ccagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     3301 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3361 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgttgg tatctcagtt
     3421 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3481 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3541 cactggcagc agccattggt aactgattta gaggactttg tcttgaagtt atgcacctgt
     3601 taaggctaaa ctgaaagaac agattttggt gagtgcggtc ctccaaccca cttaccttgg
     3661 ttcaaagagt tggtagctca gcgaaccttg agaaaaccac cgttggtagc ggtggttttt
     3721 ctttatttat gagatgatga atcaatcggt ctatcaagtc aacgaacagc tattccgtta
     3781 ctctagattt cagtgcaatt tatctcttca aatgtagcac ctgaagtcag ccccatacga
     3841 tataagttgt aattctcatg ttagtcatgc cccgcgccca ccggaaggag ctgactgggt
     3901 tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat
     3961 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     4021 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt
     4081 cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt
     4141 tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt
     4201 aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatgtcc
     4261 gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg
     4321 ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga
     4381 aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga
     4441 gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc
     4501 gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta
     4561 ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat
     4621 aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga
     4681 tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag
     4741 gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg
     4801 cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca
     4861 acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa
     4921 ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc
     4981 tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat
     5041 aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc
     5101 ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg
     5161 cgactcctgc attaggaaat taatacgact cactata
//
