LOCUS       sequence-255275-        7165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid for eGFP-(SUMO)3 client to assess 
            SUMO-SIM scaffolds..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7165)
  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 7165)
  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)3
FEATURES             Location/Qualifiers
     source          1..7165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(75..94)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     422..443
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1452..1472
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1515..1532
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1585..1603)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1599..1646
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1683..2138
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1770..1789)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1980..2001
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2164..2268
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2269..2337,2338..3129)
                     /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(2487..2506)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3300..3888
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3789..3808
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4042..4059
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4074..4213
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4139..4158)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4299..4321
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4315..4506)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5315..6397)
                     /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     6359..6378
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6398..6475)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6681..6700
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6784..6803
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6784..6802
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6803..6827
                     /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             6842..6864
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             6877..6906
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             6910..6930
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(6934..6936,6937..6939,6940..7165,1..485)
                     /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(6979..7000)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
       61 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
      121 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
      181 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
      241 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
      301 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
      361 acaaccacta cctgagcacc cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc
      421 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
      481 acaagtccgg aggctccggc gggagcggtg ggagttgggg agggtctatg tcagaagaga
      541 agcctaagga gggcgtgaaa accgagaatg accacattaa cttgaaggtt gctggacagg
      601 atgggtccgt ggttcagttc aaaattaaac gccacacacc gttgtcgaaa ctgatgaaag
      661 cgtactgcga acgccagggt ctttctatgc gtcagattcg cttccgcttt gatggtcaac
      721 cgattaatga aacggacacg ccggcacagc tggagatgga ggatgaagac actatcgacg
      781 tatttcagca gcagaccgtc gtagggggct ccggcggaag cggggggtcg ggaggttcca
      841 tgagcgaaga gaaaccaaaa gaaggagtta aaaccgaaaa cgaccatatc aacctgaagg
      901 tcgccggtca ggacggctca gtcgtgcagt tcaagattaa acgtcatacc ccattaagta
      961 agctgatgaa ggcgtattgt gagcgccaag gcttgtcgat gcgccagatt cgttttcgct
     1021 ttgatggtca gccaatcaac gaaaccgaca ccccggcgca gctggaaatg gaagatgaag
     1081 ataccatcga tgtgttccaa cagcaaacgg ttgttggcgg aagcggcggg agcggtggta
     1141 gcgggggttc tatgtccgaa gaaaaaccga aagagggtgt gaagactgaa aacgaccaca
     1201 ttaatctgaa ggtggcgggc caggacggtt ccgttgttca gtttaaaatc aaacgtcaca
     1261 ccccgctgtc gaaactgatg aaagcctatt gcgaacgtca agggctgagc atgcgtcaaa
     1321 ttcgttttcg ctttgatggt cagccaatca acgaaaccga tacaccggcg cagctggaga
     1381 tggaagacga ggacaccatt gacgtcttcc agcaacaaac tgtagtgggt ggctcctgcg
     1441 gtggctctct cgagaacctg tattttcagg gccgtaaacg taaacgtaaa cgtaaacgta
     1501 aaggctgagt cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa
     1561 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     1621 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg
     1681 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     1741 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     1801 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     1861 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     1921 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     1981 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     2041 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     2101 taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat
     2161 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     2221 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     2281 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     2341 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     2401 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     2461 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     2521 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     2581 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     2641 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     2701 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     2761 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg
     2821 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     2881 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     2941 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     3001 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     3061 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     3121 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     3181 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     3241 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     3301 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     3361 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     3421 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     3481 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     3541 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     3601 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     3661 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     3721 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     3781 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     3841 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     3901 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     3961 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     4021 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     4081 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta
     4141 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
     4201 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     4261 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     4321 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     4381 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     4441 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     4501 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
     4561 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
     4621 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
     4681 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
     4741 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
     4801 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
     4861 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
     4921 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc
     4981 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa
     5041 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc
     5101 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac
     5161 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca
     5221 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
     5281 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     5341 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     5401 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca
     5461 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa
     5521 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt
     5581 atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg
     5641 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca
     5701 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta
     5761 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg
     5821 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat
     5881 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct
     5941 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg
     6001 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat
     6061 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc
     6121 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca
     6181 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg
     6241 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt
     6301 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg
     6361 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct
     6421 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
     6481 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg
     6541 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
     6601 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
     6661 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
     6721 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga
     6781 aattaatacg actcactata ggggaattgt gagcggataa caattcccct ctagaaataa
     6841 ttttgtttaa ctttaagaag gagatatacc atgggccatc accatcacca tcaccatcac
     6901 catcacggcg aaaacctata cttccaggga tccatggtga gcaagggcga ggagctgttc
     6961 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     7021 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     7081 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     7141 cagtgcttca gccgctaccc cgacc
//
