LOCUS       sequence-449506-        5893 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent protein to reconstruct the mixture that mimics the 
            charge profile of E. coli lysate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5893)
  AUTHORS   Ahn SY, Obermeyer AC
  TITLE     Selectivity of Complex Coacervation in Multiprotein Mixtures.
  JOURNAL   JACS Au. 2024 Sep 17;4(10):3800-3812. doi: 10.1021/jacsau.4c00399. 
            eCollection 2024 Oct 28.
  PUBMED    39483238
REFERENCE   2  (bases 1 to 5893)
  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: 
            "10.1021/jacsau.4c00399"; journalName: "JACS Au"; date: "2024-09-17-
            17"; volume: "4"; issue: "10"; pages: "3800-3812"
COMMENT     Sequence Label: pET-6xHis-GFP(-12)
FEATURES             Location/Qualifiers
     source          1..5893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             456..647
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(641..663)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    749..888
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     804..823
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(903..920)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1074..1662)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1154..1173)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1743..1835
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1836..1904,1905..2696)
                     /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(2054..2073)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2811..3266)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2948..2969)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3160..3179
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(3303..3350)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     3346..3364
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(3402..3446)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(4194..4211)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             4224..4246
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4261..4285
                     /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(4285..4304)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4286..4304)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4392..4411)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4617..4694
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4695..5777
                     /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(4714..4733)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt actggttagc
       61 agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga
      121 cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga
      181 agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc aggatgctgc tggctaccct
      241 gtggaacacc tacatctgta ttaacgaagc gctggcattg accctgagtg atttttctct
      301 ggtcccgccg catccatacc gccagttgtt taccctcaca acgttccagt aaccgggcat
      361 gttcatcatc agtaacccgt atcgtgagca tcctctctcg tttcatcggt atcattaccc
      421 ccatgaacag aaatccccct tacacggagg catcagtgac caaacaggaa aaaaccgccc
      481 ttaacatggc ccgctttatc agaagccaga cattaacgct tctggagaaa ctcaacgagc
      541 tggacgcgga tgaacaggca gacatctgtg aatcgcttca cgaccacgct gatgagcttt
      601 accgcagctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
      661 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
      721 cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg
      781 gagtgtatac tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccattg
      841 cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct
      901 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
      961 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     1021 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     1081 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     1141 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     1201 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     1261 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     1321 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     1381 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     1441 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     1501 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     1561 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     1621 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     1681 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgacc
     1741 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     1801 ataaatgctt caatcatgat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     1861 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     1921 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     1981 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     2041 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     2101 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     2161 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     2221 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     2281 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     2341 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     2401 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     2461 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     2521 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     2581 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     2641 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     2701 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     2761 ggatctaggt gaagatcctt tttgataatc tcatgccatc gtgccgccag aaattgtaaa
     2821 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     2881 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     2941 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     3001 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     3061 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     3121 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     3181 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     3241 gcttaatgcg ccgctacagg gcgcgtccca ttcgccaatc cggatatagt tcctcctttc
     3301 agcaaaaaac ccctcaagac ccgtttagag gccccaaggg gttatgctag ttattgctca
     3361 gcggtggcag cagcctaggt taattaagct gcgctagtag acgagtccat gtgctggcgt
     3421 tcaaatttcg cagcagcggt ttctttacca gactcgaggg tacctcatta ttacttgtac
     3481 agctcgtcca ttccatgcgt aatgcctgca gcggtcacga attctaacag aaccatgtga
     3541 tcgcggtctt cgttcggatc tttgctcagg gctgattgcg tgctcaggta gtggttgtca
     3601 ggtaacagta caggcccatc accaatcggc gtgttttgtt gataatgatc cgctaattga
     3661 acgctgccgt cctccacgtt atggcgaatt ttaaagtttg ccttgatacc gttttcctgt
     3721 ttatcggccg taatatacac gttatgagaa ttaaagttat attccagctt atggcctaaa
     3781 atgttgccgt cttctttgaa gtcgatacct ttcagttcga tacggtttac taatgtgtcg
     3841 ccttcaaatt tcacttccgc acgcgtttta taggtgccat cgtccttgaa gctgattgta
     3901 cgctcttgaa catatccttc aggcatggct gatttgaaaa aatcgtgttg tttcatatga
     3961 tctgggtaac gactgaagca ctgaacgcca taggtcaggg tggtcactaa ggtgggccac
     4021 ggaacgggta atttgcctgt tgtgcaaata aacttcaggg tcaatttacc ctcggtagcg
     4081 tcaccttcgc cctcgccgcg cacgctaaat ttgtgaccgt tcacgtcgcc gtctaattcc
     4141 actaagatcg gtactacacc agtaaacagc tcttcacctt tgctagcgcc accgtgatgg
     4201 tggtggtgat gacccatggt atatctcctt cttaaagtta aacaaaatta tttctagagg
     4261 ggaattgtta tccgctcaca attcccctat agtgagtcgt attaatttcg cgggatcgag
     4321 atcgatctcg atcctctacg ccggacgcat cgtggccggc atcaccggcg ccacaggtgc
     4381 ggttgctggc gcctatatcg ccgacatcac cgatggggaa gatcgggctc gccacttcgg
     4441 gctcatgagc gcttgtttcg gcgtgggtat ggtggcaggc cccgtggccg ggggactgtt
     4501 gggcgccatc tccttgcatg caccattcct tgcggcggcg gtgctcaacg gcctcaacct
     4561 actactgggc tgcttcctaa tgcaggagtc gcataaggga gagcgtcgag atcccggaca
     4621 ccatcgaatg gcgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat
     4681 tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct
     4741 cttatcagac cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg
     4801 aaaaagtgga agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac
     4861 tggcgggcaa acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc
     4921 cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg
     4981 tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg
     5041 cgcaacgcgt cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg
     5101 tggaagctgc ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca
     5161 tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg
     5221 cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc
     5281 tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac
     5341 gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg
     5401 gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg
     5461 ccattaccga gtccgggctg cgcgttggtg cggacatctc ggtagtggga tacgacgata
     5521 ccgaagacag ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc
     5581 tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca
     5641 atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa
     5701 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     5761 tggaaagcgg gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcaccg
     5821 ggatctcgac cgatgccctt gagagccttc aacccagtca gctccttccg gtgggcgcgg
     5881 ggcatgacta gca
//
