LOCUS       sequence-405662-        5957 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent protein with charge-patterned cationic peptide to 
            promote biomolecular condensate formation in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5957)
  AUTHORS   Liao J, Yeong V, Obermeyer AC
  TITLE     Charge-Patterned Disordered Peptides Tune Intracellular Phase 
            Separation in Bacteria.
  JOURNAL   ACS Synth Biol. 2024 Feb 3. doi: 10.1021/acssynbio.3c00564.
  PUBMED    38308651
REFERENCE   2  (bases 1 to 5957)
  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: "ACS Synth 
            Biol. 2024 Feb 3. doi: 10.1021/acssynbio.3c00564."
COMMENT     Sequence Label: pET-6xHis-mCherry(-6)-L3
FEATURES             Location/Qualifiers
     source          1..5957
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1054..1245
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1239..1261)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1347..1486
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1402..1421
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1501..1518)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1672..2260)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1752..1771)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2341..2433
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2434..2502,2503..3294)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2652..2671)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(3409..3864)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3546..3567)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3758..3777
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(3901..3948)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     3944..3962
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(4000..4044)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(4144..4848)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(4419..4438)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     4688..4706
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(4855..4872)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             4886..4908
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4923..4947
                     /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)."
     primer_bind     complement(4947..4966)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4948..4966)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5054..5073)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        5279..5356
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5357..5957,1..482)
                     /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     complement(5376..5395)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa ggcgactgga
       61 gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc gttcccactg
      121 cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt accgagtccg
      181 ggctgcgcgt tggtgcggac atctcggtag tgggatacga cgataccgaa gacagctcat
      241 gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg caaaccagcg
      301 tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag ctgttgcccg
      361 tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc tctccccgcg
      421 cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
      481 gagcgcaacg caattaatgt aagttagctc actcattagg caccgggatc tcgaccgatg
      541 cccttgagag ccttcaaccc agtcagctcc ttccggtggg cgcggggcat gactagcatg
      601 atcgtgctcc tgtcgttgag gacccggcta ggctggcggg gttgccttac tggttagcag
      661 aatgaatcac cgatacgcga gcgaacgtga agcgactgct gctgcaaaac gtctgcgacc
      721 tgagcaacaa catgaatggt cttcggtttc cgtgtttcgt aaagtctgga aacgcggaag
      781 tcagcgccct gcaccattat gttccggatc tgcatcgcag gatgctgctg gctaccctgt
      841 ggaacaccta catctgtatt aacgaagcgc tggcattgac cctgagtgat ttttctctgg
      901 tcccgccgca tccataccgc cagttgttta ccctcacaac gttccagtaa ccgggcatgt
      961 tcatcatcag taacccgtat cgtgagcatc ctctctcgtt tcatcggtat cattaccccc
     1021 atgaacagaa atccccctta cacggaggca tcagtgacca aacaggaaaa aaccgccctt
     1081 aacatggccc gctttatcag aagccagaca ttaacgcttc tggagaaact caacgagctg
     1141 gacgcggatg aacaggcaga catctgtgaa tcgcttcacg accacgctga tgagctttac
     1201 cgcagctgcc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     1261 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     1321 tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga
     1381 gtgtatactg gcttaactat gcggcatcag agcagattgt actgagagtg caccattgcg
     1441 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgct cttccgcttc
     1501 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     1561 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     1621 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1681 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1741 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1801 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1861 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1921 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1981 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     2041 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     2101 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     2161 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     2221 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     2281 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgaccta
     2341 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     2401 aaatgcttca atcatgattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     2461 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     2521 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     2581 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     2641 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     2701 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     2761 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     2821 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     2881 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     2941 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     3001 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     3061 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     3121 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     3181 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     3241 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     3301 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     3361 atctaggtga agatcctttt tgataatctc atgccatcgt gccgccagaa attgtaaacg
     3421 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     3481 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     3541 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     3601 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     3661 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     3721 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     3781 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     3841 ttaatgcgcc gctacagggc gcgtcccatt cgccaatccg gatatagttc ctcctttcag
     3901 caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctagtt attgctcagc
     3961 ggtggcagca gcctaggtta attaagctgc gctagtagac gagtccatgt gctggcgttc
     4021 aaatttcgca gcagcggttt ctttaccaga ctcgagttat tacttccgcg atttcttccc
     4081 tcttcgaccc ttccgcgatt tcttccctct tcgacccttt cgagatttct tgcctcttcg
     4141 tcccttgtac agctcgtcca tgccgccggt ggagtggcgg ccctcggcgc gttcgtactg
     4201 ttccacgatg gtgtagtcct cgttgtggga ggtgatgtcc aacttgatgt tgacgttgta
     4261 ggcgccgggc agctgcacgg gcttcttggc cttgtaggtg gtcttgacct cagcgtcgta
     4321 gtggccgccg tccttcagct tcagcctctg cttgatctcg cccttcaggg cgccgtcctc
     4381 ggggtacatc cgctcggagg aggcctccca gcccatggtc ttcttctgca ttacggggcc
     4441 gtcggagggg aagttggtgc cgcgcagctt caccttgtag atgaactcgc cgtcctggag
     4501 ggaggagtcc tgggtcacgg tcaccacgcc gccgtcctcg aagttcatca cgcgctccca
     4561 cttgaagccc tcggggaagg acagcttcaa gtagtcgggg atgtcggcgg ggtgcttcac
     4621 gtaggccttg gagccgtaca tgaactgagg ggacaggatg tcccaggcga agggcagggg
     4681 gccacccttg gtcaccttca gcttggcggt ctgggtgccc tcgtaggggc ggccctcgcc
     4741 ctcgccctcg atctcgaact cgtggccgtt cacggagccc tccatgtgca ccttgaagcg
     4801 catgaactcc ttgatgatgg ccatgttatc ctcctcgccc ttgctcacgc caccgtgatg
     4861 gtggtggtga tgacccatat gtatatctcc ttcttaaagt taaacaaaat tatttctaga
     4921 ggggaattgt tatccgctca caattcccct atagtgagtc gtattaattt cgcgggatcg
     4981 agatcgatct cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt
     5041 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc
     5101 gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg
     5161 ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac
     5221 ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga
     5281 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
     5341 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
     5401 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     5461 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     5521 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     5581 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     5641 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     5701 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     5761 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     5821 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     5881 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     5941 tctgcgtctg gctggct
//
