LOCUS       sequence-255197-        6541 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid for eGFP-(SUMO)1 client to assess 
            SUMO-SIM scaffolds..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6541)
  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 6541)
  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-(SUMO)1
FEATURES             Location/Qualifiers
     source          1..6541
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /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             40..62
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             75..104
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             108..128
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(132..134,135..137,138..848)
                     /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(177..198)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(438..457)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     785..806
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1191..1211
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1254..1271
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1324..1342)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1338..1385
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1422..1877
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1509..1528)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1719..1740
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1903..2007
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2008..2076,2077..2868)
                     /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(2226..2245)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3039..3627
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3528..3547
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3781..3798
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3813..3952
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3878..3897)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4038..4060
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4054..4245)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5054..6136)
                     /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     6098..6117
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6137..6214)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6420..6439
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(6523..6541,1)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6523..6541
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
       61 gatataccat gggccatcac catcaccatc accatcacca tcacggcgaa aacctatact
      121 tccagggatc catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
      181 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
      241 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
      301 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
      361 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
      421 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
      481 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
      541 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
      601 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
      661 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
      721 ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac gagaagcgcg
      781 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
      841 tgtacaagtc cggaggctcc ggcgggagcg gtgggagttg gggagggtct atgtcagaag
      901 agaagcctaa ggagggcgtg aaaaccgaga atgaccacat taacttgaag gttgctggac
      961 aggatgggtc cgtggttcag ttcaaaatta aacgccacac accgttgtcg aaactgatga
     1021 aagcgtactg cgaacgccag ggtctttcta tgcgtcagat tcgcttccgc tttgatggtc
     1081 aaccgattaa tgaaacggac acgccggcac agctggagat ggaggatgaa gacactatcg
     1141 acgtatttca gcagcagacc gtcgtagggg gctcctgcgg cggaagcctc gagaacctgt
     1201 attttcaggg ccgtaaacgt aaacgtaaac gtaaacgtaa aggctgagtc gagcaccacc
     1261 accaccacca ctgagatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg
     1321 ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt
     1381 ttttgctgaa aggaggaact atatccggat tggcgaatgg gacgcgccct gtagcggcgc
     1441 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     1501 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     1561 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     1621 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     1681 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     1741 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     1801 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     1861 attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1921 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1981 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     2041 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     2101 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     2161 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     2221 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     2281 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     2341 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     2401 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     2461 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     2521 ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta
     2581 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     2641 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     2701 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     2761 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     2821 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     2881 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2941 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     3001 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     3061 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3121 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3181 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     3241 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3301 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3361 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3421 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     3481 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     3541 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     3601 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     3661 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     3721 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     3781 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     3841 ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat
     3901 agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca
     3961 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     4021 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     4081 acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc
     4141 tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct
     4201 gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta
     4261 agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat
     4321 acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc
     4381 ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa
     4441 tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat
     4501 aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac
     4561 cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc
     4621 gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct
     4681 caacgacagg agcacgatca tgcgcacccg tggggccgcc atgccggcga taatggcctg
     4741 cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa
     4801 gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa agcggtcctc
     4861 gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga taaagaagac
     4921 agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt
     4981 gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt
     5041 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5101 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc
     5161 ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt
     5221 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta
     5281 acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatatccg
     5341 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt
     5401 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa
     5461 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag
     5521 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg
     5581 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac
     5641 cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata
     5701 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat
     5761 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg
     5821 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc
     5881 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa
     5941 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat
     6001 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct
     6061 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata
     6121 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca
     6181 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc
     6241 gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca
     6301 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
     6361 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     6421 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     6481 atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag
     6541 g
//
