LOCUS       sequence-255272-        6334 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid for eGFP-(SIM)1 client to assess 
            SUMO-SIM scaffolds..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6334)
  AUTHORS   Banani SF, Rice AM, Peeples WB, Lin Y, Jain S, Parker R, Rosen MK
  TITLE     Compositional Control of Phase-Separated Cellular Bodies.
  JOURNAL   Cell. 2016 Jul 28;166(3):651-663. doi: 10.1016/j.cell.2016.06.010. 
            Epub 2016 Jun 30.
  PUBMED    27374333
REFERENCE   2  (bases 1 to 6334)
  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"; doi: 
            "10.1016/j.cell.2016.06.010"; journalName: "Cell"; date: 
            "2016-07-28- 28"; volume: "166"; issue: "3"; pages: "651-663"
COMMENT     Sequence Label: GFP-(SIM)1
FEATURES             Location/Qualifiers
     source          1..6334
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             94..123
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             127..147
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(151..153,154..156,157..867)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(196..217)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(457..476)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     804..825
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1003..1023
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1066..1083
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1136..1154)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1150..1197
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1234..1689
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1321..1340)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1531..1552
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1715..1819
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1820..1888,1889..2680)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2038..2057)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2851..3439
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3340..3359
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3593..3610
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3625..3764
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3690..3709)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3850..3872
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3866..4057)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4866..5948)
                     /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     5910..5929
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5949..6026)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6232..6251
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggccatcacc atcaccatca ccatcaccat
      121 cacggcgaaa acctatactt ccagggatcc atggtgagca agggcgagga gctgttcacc
      181 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
      241 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
      301 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
      361 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
      421 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
      481 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
      541 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
      601 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
      661 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
      721 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa gctgagcaaa
      781 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
      841 actctcggca tggacgagct gtacaagtcc ggaggtagtg gcggtagcgg tggctcttgg
      901 ggtgggagca aagtagatgt gatcgacttg accatcgaaa gcagcagcga cgaagaagaa
      961 gatccgcctg ccaaacgcgg cggtagctgc ggtggctctc tcgagaacct gtattttcag
     1021 ggccgtaaac gtaaacgtaa acgtaaacgt aaaggctgag tcgagcacca ccaccaccac
     1081 cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct
     1141 gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg
     1201 aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     1261 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     1321 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     1381 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     1441 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     1501 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     1561 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     1621 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     1681 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     1741 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     1801 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     1861 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     1921 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     1981 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     2041 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     2101 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     2161 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     2221 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     2281 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     2341 cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg
     2401 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     2461 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     2521 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     2581 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     2641 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     2701 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     2761 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     2821 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     2881 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     2941 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     3001 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     3061 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     3121 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     3181 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     3241 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     3301 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     3361 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     3421 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     3481 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     3541 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     3601 agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg
     3661 tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     3721 cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa
     3781 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     3841 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     3901 ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt
     3961 catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc
     4021 gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat
     4081 ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta
     4141 ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga
     4201 tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg
     4261 taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc
     4321 agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg
     4381 ttgttgctca ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg
     4441 gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca
     4501 ggagcacgat catgcgcacc cgtggggccg ccatgccggc gataatggcc tgcttctcgc
     4561 cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga
     4621 ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa
     4681 tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa
     4741 gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc
     4801 tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt
     4861 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4921 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc
     4981 agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag
     5041 cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg
     5101 atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg
     5161 cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc
     5221 agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac
     5281 atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat
     5341 ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc
     5401 gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca
     5461 tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga
     5521 acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg
     5581 atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg
     5641 ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta
     5701 atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc
     5761 agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc
     5821 gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc
     5881 acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact
     5941 ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga
     6001 aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat gcgactcctg
     6061 cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg
     6121 tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac
     6181 gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc
     6241 ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc
     6301 ggcgtagagg atcgagatct cgatcccgcg aaat
//
