LOCUS       sequence-449507-        5970 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 5970)
  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 5970)
  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: pET28-a(+)-6xHis-mBlueberry(-9)
FEATURES             Location/Qualifiers
     source          1..5970
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     374..393
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     458..477
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      588..1176
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1077..1096
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1330..1347
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1362..1501
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1427..1446)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1587..1609
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1603..1794)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2603..3685)
                     /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     3647..3666
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3686..3763)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3969..3988
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4072..4091
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4072..4090
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4091..4115
                     /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             4130..4152
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4165..4182
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4906..4923
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4976..4994)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4990..5037
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      5074..5529
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5161..5180)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5371..5392
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(5621..5970,1..466))
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(join(5621..5970,1..466))
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
ORIGIN
        1 ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct gttaaaagga caattacaaa
       61 caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg
      121 aatcaggata ttcttctaat acctggaatg ctgttttccc ggggatcgca gtggtgagta
      181 accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg
      241 tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat
      301 gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg
      361 attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat
      421 ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg gctcataaca ccccttgtat
      481 tactgtttat gtaagcagac agttttattg ttcatgacca aaatccctta acgtgagttt
      541 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
      601 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
      661 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
      721 ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta
      781 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
      841 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
      901 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
      961 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     1021 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     1081 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     1141 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     1201 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     1261 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     1321 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc
     1381 cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca atctgctctg
     1441 atgccgcata gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc
     1501 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     1561 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     1621 atcaccgaaa cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc
     1681 acagatgtct gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt
     1741 ctggcttctg ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc
     1801 ctccgtgtaa gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat
     1861 gctcacgata cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa
     1921 acaactggcg gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg
     1981 cttcgttaat acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat
     2041 ccggaacata atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa
     2101 accgaagacc attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca
     2161 cgttcgctcg cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag
     2221 ccgggtcctc aacgacagga gcacgatcat gcgcacccgt ggggccgcca tgccggcgat
     2281 aatggcctgc ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag
     2341 ggcgtgcaag attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa
     2401 gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat
     2461 aaagaagaca gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct
     2521 gactgggttg aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta
     2581 acttacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     2641 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg
     2701 tggtttttct tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct
     2761 gagagagttg cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga
     2821 tggtggttaa cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg
     2881 agatatccgc accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca
     2941 tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg
     3001 tttgttgaaa accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt
     3061 gattgcgagt gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta
     3121 atgggcccgc taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca
     3181 gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat
     3241 caagaaataa cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat
     3301 ccagcggata gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg
     3361 ctttacaggc ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt
     3421 gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg
     3481 aggtggcaac gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg
     3541 gaatgtaatt cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt
     3601 ggctggcctg gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga
     3661 catcgtataa cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct
     3721 atcatgccat accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct
     3781 cccttatgcg actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc
     3841 gccgccgcaa ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg
     3901 cctgccacca tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct
     3961 tccccatcgg tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg
     4021 ccggccacga tgcgtccggc gtagaggatc gagatctcga tcccgcgaaa ttaatacgac
     4081 tcactatagg ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact
     4141 ttaagaagga gatataccat gggccatcat catcatcatc atggcggcgt gagcaaaggc
     4201 gaagaaaaca acgtggcgat tattaaagaa tttatgcgct ttaaagtgca tatggaaggc
     4261 agcgtgaacg gccatgaatt tgaaattgaa ggcgaaggcg aaggccgccc gtatgaaggc
     4321 acccagaccg cgaaactgaa agtgaccaaa ggcggcccgc tgccgtttgc gtgggatatt
     4381 ctgagcccgc agtttctgtt tggcagcaaa gtgtatatta aacatccggc ggatattccg
     4441 gattatttta aactgagctt tccggaaggc tttaaatggg aacgcgtgat gaactttgaa
     4501 gatggcggcg tggtgaccgt gacccaggat agcagcctgc aggatggcgt gtttatttat
     4561 aaagtggaac tgcgcggcac caactttccg agcgatggcc cggtgatgca gaaaaaaacc
     4621 atgggctggg aagcgtttag cgaacgcatg tatccggaag atggcgcgct gaaaagcgaa
     4681 attaaaaccc gcctgaaact gaaagatggc ggccattatg atgcggaagt gaaaaccacc
     4741 tataaagcga aagagccggt gcagctgccg ggcgcgtata acgtgaacat taaactggat
     4801 attgtgagcc ataacgaaga ttataccatt gtggaacagt atgaacgcgc ggaaggcgaa
     4861 catagcaccg gcggcatgga tgaactgtat aaataataac tcgagcacca ccaccaccac
     4921 cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct
     4981 gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg
     5041 aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     5101 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     5161 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     5221 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     5281 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     5341 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     5401 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     5461 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     5521 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     5581 ctaaatacat tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca
     5641 tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc
     5701 gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt
     5761 atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa
     5821 aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca
     5881 aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa
     5941 aatcactcgc atcaaccaaa ccgttattca
//
