LOCUS       sequence-245480-        5123 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of GFP-SZ2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5123)
  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 5123)
  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_GFP-SZ2
FEATURES             Location/Qualifiers
     source          1..5123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     46..65
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     149..168
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        149..167
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    168..192
                     /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             207..229
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             240..257
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(258..260,261..971)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(300..321)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(561..580)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     908..929
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             984..1004
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1167..1184
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1237..1255)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1251..1298
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1335..1790
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1422..1441)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1632..1653
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1905..2720)
                     /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(1905..2720)
                     /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     2628..2647
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2712..2731
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2873..3461
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3362..3381
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3615..3632
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3803..4885)
                     /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     4847..4866
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4886..4963)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
       61 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      121 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      181 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat atacatatgc
      241 atcaccacca tcatcatgtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
      301 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
      361 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
      421 tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc
      481 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
      541 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
      601 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
      661 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg
      721 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
      781 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
      841 tgcccgacaa ccactacctg agcacccagt ccaagctgag caaagacccc aacgagaagc
      901 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
      961 agctgtacaa gggcggcgga tctgagaatt tatatttcca gggcgctcgt aatgcatacc
     1021 tgcgtaaaaa aatcgcgcgt ttaaaaaaag acaacttgca acttgagcgc gacgaacaaa
     1081 atttggagaa aataatcgcc aaccttcggg acgagatcgc gcgtctggaa aacgaggtgg
     1141 cttcgcatga gcaataatga ctcgagcacc accaccacca ccactgagat ccggctgcta
     1201 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     1261 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
     1321 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1381 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1441 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1501 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1561 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1621 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     1681 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     1741 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact
     1801 tttcggggta cccaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     1861 ttttctaaat acattcaaat atgtatccgc tcatgaatta attcttagaa aaactcatcg
     1921 agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa
     1981 agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc
     2041 tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg
     2101 tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat
     2161 ggcaaaagtt tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca
     2221 tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga
     2281 aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg
     2341 aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg
     2401 aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata
     2461 aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca
     2521 tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg
     2581 ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat
     2641 ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctaga gcaagacgtt
     2701 tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt
     2761 attgttcatg accaaaatcc cttaacgtga cctaggtcat gaccaaaatc ccttaacgtg
     2821 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     2881 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     2941 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     3001 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     3061 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     3121 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     3181 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     3241 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     3301 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     3361 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     3421 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     3481 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     3541 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     3601 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccat aatccatgat
     3661 aaagaagaca gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct
     3721 gactgggttg aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta
     3781 acttacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     3841 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg
     3901 tggtttttct tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct
     3961 gagagagttg cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga
     4021 tggtggttaa cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg
     4081 agatatccgc accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca
     4141 tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg
     4201 tttgttgaaa accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt
     4261 gattgcgagt gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta
     4321 atgggcccgc taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca
     4381 gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat
     4441 caagaaataa cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat
     4501 ccagcggata gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg
     4561 ctttacaggc ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt
     4621 gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg
     4681 aggtggcaac gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg
     4741 gaatgtaatt cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt
     4801 ggctggcctg gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga
     4861 catcgtataa cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct
     4921 atcatgccat accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct
     4981 cccttatgcg actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc
     5041 gccgccgcaa ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg
     5101 cctgccacca tacccacgcc gaa
//
