LOCUS       sequence-288551-        5539 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsRck_296-472-HsPatL1_1-173.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5539)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 5539)
  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_1-173 (MCSII)_D
FEATURES             Location/Qualifiers
     source          1..5539
                     /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             2076..2120
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2158..2176)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2172..2219
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2449..3264)
                     /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(2449..3264)
                     /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     3172..3191
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3265..3356)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3372..4121)
                     /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(3382..3399)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3622..3641)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4317..5399)
                     /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     5361..5380
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5400..5477)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(5523..5539,1..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 agctaccagt ggcagttaat gaacaaacag
     1741 gcaatggaga gagagatgaa atggacttgt tgggtgacca tgaggagaat ctggcagaaa
     1801 ggctcagtaa gatggtgatt gaaaatgaac tagaagatcc agctattatg agggcagtgc
     1861 agaccaggcc agttttacaa ccccaaccag gaagtctgaa ttccagtatc tgggatggat
     1921 ctgaagttct gaggcgaatc cgaggaccac tgcttgctca ggaaatgcct acagtgtctg
     1981 tattagaata tgctttgcct cagaggcccc cccagggtcc agaagatgat cgggaccttt
     2041 ctgaacgagc attatagggt accctcgagt ctggtaaaga aaccgctgct gcgaaatttg
     2101 aacgccagca catggactcg tctactagcg cagcttaatt aacctaggct gctgccaccg
     2161 ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     2221 tgaaacctca ggcatttgag aagcacacgg tcacactgct tccggtagtc aataaaccgg
     2281 taaaccagca atagacataa gcggctattt aacgaccctg ccctgaaccg acgacaagct
     2341 gacgaccggg tctccgcaag tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     2401 ttatttttct aaatacattc aaatatgtat ccgctcatga attaattctt agaaaaactc
     2461 atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg
     2521 aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag
     2581 atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc
     2641 ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga
     2701 gaatggcaaa agtttatgca tttctttcca gacttgttca acaggccagc cattacgctc
     2761 gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag
     2821 acgaaatacg cggtcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg
     2881 caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac
     2941 ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg
     3001 gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat
     3061 ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc
     3121 atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc
     3181 ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc tagagcaaga
     3241 cgtttcccgt tgaatatggc tcatactctt cctttttcaa tattattgaa gcatttatca
     3301 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     3361 catgcagcgc tcttccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg
     3421 gcgagcggtg tcagctcact caaaagcggt aatacggtta tccacagaat caggggataa
     3481 agccggaaag aacatgtgag caaaaagcaa agcaccggaa gaagccaacg ccgcaggcgt
     3541 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agccagaggt
     3601 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     3661 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     3721 gcgtggcgct ttctcatagc tcacgctgtt ggtatctcag ttcggtgtag gtcgttcgct
     3781 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     3841 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccattg
     3901 gtaactgatt tagaggactt tgtcttgaag ttatgcacct gttaaggcta aactgaaaga
     3961 acagattttg gtgagtgcgg tcctccaacc cacttacctt ggttcaaaga gttggtagct
     4021 cagcgaacct tgagaaaacc accgttggta gcggtggttt ttctttattt atgagatgat
     4081 gaatcaatcg gtctatcaag tcaacgaaca gctattccgt tactctagat ttcagtgcaa
     4141 tttatctctt caaatgtagc acctgaagtc agccccatac gatataagtt gtaattctca
     4201 tgttagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca
     4261 tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac
     4321 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     4381 cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg
     4441 ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg
     4501 ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat
     4561 gagctgtctt cggtatcgtc gtatcccact accgagatgt ccgcaccaac gcgcagcccg
     4621 gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca
     4681 gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc
     4741 cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag
     4801 ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc
     4861 tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa
     4921 ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg
     4981 caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca
     5041 ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt
     5101 tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg
     5161 acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac
     5221 tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc
     5281 gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa
     5341 acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca
     5401 ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg
     5461 cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa
     5521 attaatacga ctcactata
//
