LOCUS       sequence-405660-        5903 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 5903)
  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 5903)
  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)-L1
FEATURES             Location/Qualifiers
     source          1..5903
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1207..1398
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1392..1414)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1500..1639
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1555..1574
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1654..1671)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1825..2413)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1905..1924)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2494..2586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2587..2655,2656..3447)
                     /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(2805..2824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(3562..4017)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3699..3720)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3911..3930
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(4054..4101)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     4097..4115
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(4153..4197)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(4243..4947)
                     /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(4518..4537)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     4787..4805
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(4954..4971)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             4985..5007
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    5022..5046
                     /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(5046..5065)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5047..5065)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5153..5172)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        5378..5455
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5456..5903,1..635)
                     /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(5475..5494)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg
       61 gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa
      121 gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa
      181 ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca
      241 tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg
      301 cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gacatctcgg
      361 tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca
      421 aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg
      481 gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc
      541 tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      601 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtaagttag
      661 ctcactcatt aggcaccggg atctcgaccg atgcccttga gagccttcaa cccagtcagc
      721 tccttccggt gggcgcgggg catgactagc atgatcgtgc tcctgtcgtt gaggacccgg
      781 ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg
      841 tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt
      901 ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg
      961 atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag
     1021 cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt
     1081 ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc
     1141 atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag
     1201 gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag
     1261 acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt
     1321 gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat
     1381 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     1441 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     1501 gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat
     1561 cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag
     1621 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     1681 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     1741 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     1801 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     1861 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     1921 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     1981 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     2041 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     2101 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     2161 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     2221 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     2281 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     2341 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     2401 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     2461 aaactcacgt taagggattt tggtcatgac ctatttgttt atttttctaa atacattcaa
     2521 atatgtatcc gctcatgaga caataaccct gataaatgct tcaatcatga ttgaaaaagg
     2581 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     2641 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     2701 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     2761 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     2821 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     2881 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     2941 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     3001 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     3061 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     3121 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     3181 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     3241 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     3301 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     3361 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     3421 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     3481 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     3541 ctcatgccat cgtgccgcca gaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa
     3601 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     3661 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     3721 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     3781 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     3841 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     3901 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     3961 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
     4021 attcgccaat ccggatatag ttcctccttt cagcaaaaaa cccctcaaga cccgtttaga
     4081 ggccccaagg ggttatgcta gttattgctc agcggtggca gcagcctagg ttaattaagc
     4141 tgcgctagta gacgagtcca tgtgctggcg ttcaaatttc gcagcagcgg tttctttacc
     4201 agactcgagt cattactttc gagatttctt gcctcttcgt cccttgtaca gctcgtccat
     4261 gccgccggtg gagtggcggc cctcggcgcg ttcgtactgt tccacgatgg tgtagtcctc
     4321 gttgtgggag gtgatgtcca acttgatgtt gacgttgtag gcgccgggca gctgcacggg
     4381 cttcttggcc ttgtaggtgg tcttgacctc agcgtcgtag tggccgccgt ccttcagctt
     4441 cagcctctgc ttgatctcgc ccttcagggc gccgtcctcg gggtacatcc gctcggagga
     4501 ggcctcccag cccatggtct tcttctgcat tacggggccg tcggagggga agttggtgcc
     4561 gcgcagcttc accttgtaga tgaactcgcc gtcctggagg gaggagtcct gggtcacggt
     4621 caccacgccg ccgtcctcga agttcatcac gcgctcccac ttgaagccct cggggaagga
     4681 cagcttcaag tagtcgggga tgtcggcggg gtgcttcacg taggccttgg agccgtacat
     4741 gaactgaggg gacaggatgt cccaggcgaa gggcaggggg ccacccttgg tcaccttcag
     4801 cttggcggtc tgggtgccct cgtaggggcg gccctcgccc tcgccctcga tctcgaactc
     4861 gtggccgttc acggagccct ccatgtgcac cttgaagcgc atgaactcct tgatgatggc
     4921 catgttatcc tcctcgccct tgctcacgcc accgtgatgg tggtggtgat gacccatatg
     4981 tatatctcct tcttaaagtt aaacaaaatt atttctagag gggaattgtt atccgctcac
     5041 aattccccta tagtgagtcg tattaatttc gcgggatcga gatcgatctc gatcctctac
     5101 gccggacgca tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc
     5161 gccgacatca ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc
     5221 ggcgtgggta tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat
     5281 gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta
     5341 atgcaggagt cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac
     5401 ctttcgcggt atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa
     5461 accagtaacg ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg
     5521 cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat
     5581 ggcggagctg aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt
     5641 gctgattggc gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc
     5701 gattaaatct cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag
     5761 cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct
     5821 gatcattaac tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa
     5881 tgttccggcg ttatttcttg atg
//
