LOCUS       sequence-245499-        5816 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of RGG-RGG-RGG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5816)
  AUTHORS   Schuster BS, Reed EH, Parthasarathy R, Jahnke CN, Caldwell RM, 
            Bermudez JG, Ramage H, Good MC, Hammer DA
  TITLE     Controllable protein phase separation and modular recruitment to 
            form responsive membraneless organelles.
  JOURNAL   Nat Commun. 2018 Jul 30;9(1):2985. doi: 10.1038/s41467-018-05403-1.
  PUBMED    30061688
REFERENCE   2  (bases 1 to 5816)
  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"; journalName: "Nat 
            Commun."; date: "2018-07-30"; volume: "9(1):2985. doi"; pages: " 
            10.1038/s41467-018-05403-1"
COMMENT     Sequence Label: pET_RGG-RGG-RGG
FEATURES             Location/Qualifiers
     source          1..5816
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     88..107
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        88..106
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    107..131
                     /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             146..168
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             686..706
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             713..730
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1241..1264
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1271..1288
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1799..1816
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1869..1887)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1883..1930
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1967..2422
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2054..2073)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2264..2285
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2537..3352)
                     /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(2537..3352)
                     /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     3260..3279
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3344..3363
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3505..4093
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3994..4013
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4247..4264
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(4435..5517)
                     /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     5479..5498
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5518..5595)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     join(5801..5816,1..4)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag
       61 aggatcgaga tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg
      121 ataacaattc ccctctagaa ataattttgt ttaactttaa gaaggagata tacatatgga
      181 atccaatcaa tccaataacg gtggctcggg taatgccgct ctgaatcgtg gtggccgcta
      241 cgtgccgccg catctgcgtg gtggcgatgg tggtgcagca gcagctgcat ctgctggcgg
      301 tgatgaccgt cgcggcggtg cgggcggtgg cggttatcgt cgcggcggtg gcaatagtgg
      361 tggcggtggc ggtggcggtt atgatcgtgg ttacaatgac aaccgtgatg accgcgataa
      421 ccgtggcggt tccggcggtt acggtcgtga tcgtaattat gaagaccgtg gttacaacgg
      481 cggtggcggt ggcggtggca atcgtggcta taacaataac cgtggtggcg gtggcggtgg
      541 ctacaatcgc caggatcgtg gtgacggtgg cagctctaac ttttcgcgcg gtggctataa
      601 taaccgtgat gaaggcagcg acaaccgcgg tagtggccgt tcctacaata acgatcgtcg
      661 cgacaatggt ggcgatggcg aattcgaaaa tttgtatttc caaggcggat ccttggtgcc
      721 tagaggcagc atggaatcga atcaatctaa caacggtgga tcgggaaacg cagcattgaa
      781 tcgtgggggc cgttacgtac caccacactt gagaggcgga gatggtggtg ccgccgctgc
      841 tgcctcagcc ggaggcgacg acagacgcgg tggggccggg ggtggtggct atcgtagagg
      901 tggtggaaat tcagggggcg gcggcggagg aggctacgat cgtggataca acgacaaccg
      961 ggatgaccgt gacaatcggg gcggctccgg cggttatggc cgggacagaa attacgagga
     1021 tcgcggttat aacgggggtg gtgggggagg aggcaatcgc ggctacaata ataaccgcgg
     1081 tgggggtgga ggcggctata atcggcagga ccgtggagac ggcggttcta gtaatttctc
     1141 gcggggaggg tataacaaca gagatgaggg ctcagacaac agaggctcag gcagatcata
     1201 caacaacgat cggcgcgaca atggtggtga cgggggtacc ctggaggtgc tttttcaggg
     1261 ccctgaattc ttagtacctc gcgggtcaat ggaatccaat caatccaata acggtggctc
     1321 gggtaatgcc gctctgaatc gtggtggccg ctacgtgccg ccgcatctgc gtggtggcga
     1381 tggtggtgca gcagcagctg catctgctgg cggtgatgac cgtcgcggcg gtgcgggcgg
     1441 tggcggttat cgtcgcggcg gtggcaatag tggtggcggt ggcggtggcg gttatgatcg
     1501 tggttacaat gacaaccgtg atgaccgcga taaccgtggc ggttccggcg gttacggtcg
     1561 tgatcgtaat tatgaagacc gtggttacaa cggcggtggc ggtggcggtg gcaatcgtgg
     1621 ctataacaat aaccgtggtg gcggtggcgg tggctacaat cgccaggatc gtggtgacgg
     1681 tggcagctct aacttttcgc gcggtggcta taataaccgt gatgaaggca gcgacaaccg
     1741 cggtagtggc cgttcctaca ataacgatcg tcgcgacaat ggtggcgatg gcctcgagca
     1801 ccaccaccac caccactgag atccggctgc taacaaagcc cgaaaggaag ctgagttggc
     1861 tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag
     1921 gggttttttg ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc
     1981 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2041 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2101 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2161 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2221 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2281 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2341 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     2401 aaaatattaa cgcttacaat ttaggtggca cttttcgggg tacccaggtg gcacttttcg
     2461 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     2521 gctcatgaat taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca
     2581 tatcaggatt atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact
     2641 caccgaggca gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc
     2701 caacatcaat acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat
     2761 caccatgagt gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga
     2821 cttgttcaac aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt
     2881 tattcattcg tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat
     2941 tacaaacagg aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt
     3001 cacctgaatc aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg
     3061 tgagtaacca tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa
     3121 attccgtcag ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt
     3181 tgccatgttt cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg
     3241 cacctgattg cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt
     3301 tggaatttaa tcgcggccta gagcaagacg tttcccgttg aatatggctc ataacacccc
     3361 ttgtattact gtttatgtaa gcagacagtt ttattgttca tgaccaaaat cccttaacgt
     3421 gacctaggtc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     3481 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     3541 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     3601 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     3661 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     3721 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     3781 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     3841 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     3901 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3961 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     4021 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     4081 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     4141 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     4201 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     4261 cgaggaagcg gaagagcgcc ataatccatg ataaagaaga cagtcataag tgcggcgacg
     4321 atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc
     4381 ggtcgagatc ccggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg
     4441 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     4501 gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg
     4561 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
     4621 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga
     4681 gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga
     4741 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
     4801 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
     4861 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
     4921 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg
     4981 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
     5041 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
     5101 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
     5161 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
     5221 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
     5281 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
     5341 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
     5401 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
     5461 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
     5521 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg
     5581 ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc
     5641 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg
     5701 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
     5761 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgata
//
