LOCUS       sequence-288550-        5353 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsRck_296-472-HsPatL1_1-111.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5353)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 5353)
  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_1-111 (MCSII)_D
FEATURES             Location/Qualifiers
     source          1..5353
                     /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             1890..1934
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1972..1990)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1986..2033
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2263..3078)
                     /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(2263..3078)
                     /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     2986..3005
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3079..3170)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3186..3935)
                     /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(3196..3213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3436..3455)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4131..5213)
                     /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     5175..5194
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5214..5291)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(5337..5353,1..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 agaccaggta gggtaccctc gagtctggta aagaaaccgc tgctgcgaaa tttgaacgcc
     1921 agcacatgga ctcgtctact agcgcagctt aattaaccta ggctgctgcc accgctgagc
     1981 aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaac
     2041 ctcaggcatt tgagaagcac acggtcacac tgcttccggt agtcaataaa ccggtaaacc
     2101 agcaatagac ataagcggct atttaacgac cctgccctga accgacgaca agctgacgac
     2161 cgggtctccg caagtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     2221 ttctaaatac attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag
     2281 catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag
     2341 ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg
     2401 gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc
     2461 aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg
     2521 caaaagttta tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc
     2581 aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa
     2641 tacgcggtcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa
     2701 cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa
     2761 tgctgttttc ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa
     2821 atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc
     2881 tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg
     2941 cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt
     3001 atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc
     3061 ccgttgaata tggctcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     3121 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggcatgca
     3181 gcgctcttcc gcttcctcgc tcactgactc gctacgctcg gtcgttcgac tgcggcgagc
     3241 ggtgtcagct cactcaaaag cggtaatacg gttatccaca gaatcagggg ataaagccgg
     3301 aaagaacatg tgagcaaaaa gcaaagcacc ggaagaagcc aacgccgcag gcgtttttcc
     3361 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagccag aggtggcgaa
     3421 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     3481 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     3541 cgctttctca tagctcacgc tgttggtatc tcagttcggt gtaggtcgtt cgctccaagc
     3601 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     3661 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc attggtaact
     3721 gatttagagg actttgtctt gaagttatgc acctgttaag gctaaactga aagaacagat
     3781 tttggtgagt gcggtcctcc aacccactta ccttggttca aagagttggt agctcagcga
     3841 accttgagaa aaccaccgtt ggtagcggtg gtttttcttt atttatgaga tgatgaatca
     3901 atcggtctat caagtcaacg aacagctatt ccgttactct agatttcagt gcaatttatc
     3961 tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgttag
     4021 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
     4081 gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg
     4141 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4201 gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac
     4261 agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt
     4321 tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg
     4381 tcttcggtat cgtcgtatcc cactaccgag atgtccgcac caacgcgcag cccggactcg
     4441 gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga
     4501 acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg
     4561 ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc
     4621 agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga
     4681 cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata
     4741 ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca
     4801 gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg
     4861 cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc
     4921 atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt
     4981 tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg
     5041 cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc
     5101 actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc
     5161 tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc
     5221 accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat
     5281 tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaaattaat
     5341 acgactcact ata
//
