LOCUS       sequence-449505-        5902 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 5902)
  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 5902)
  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(-7)-neg_tag1
FEATURES             Location/Qualifiers
     source          1..5902
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     878..897
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(917..994)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1200..1219
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1307..1326
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1307..1325
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1326..1350
                     /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             1365..1387
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1400..1417
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2174..2218
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2256..2274)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2270..2317
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2354..2809
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2441..2460)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2651..2672
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2924..3715,3716..3784))
                     /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     3547..3566
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3785..3877)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3958..4546
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4447..4466
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4700..4717
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4732..4871
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4797..4816)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4957..4979
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4973..5164)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5736..5902,1..916))
                     /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"
ORIGIN
        1 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta
       61 acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatgtccg
      121 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt
      181 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa
      241 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag
      301 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg
      361 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac
      421 cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata
      481 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat
      541 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg
      601 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc
      661 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa
      721 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat
      781 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct
      841 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata
      901 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca
      961 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc
     1021 gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca
     1081 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
     1141 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     1201 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     1261 atgcgtccgg cgtagaggat cgagatcgat ctcgatcccg cgaaattaat acgactcact
     1321 ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag
     1381 aaggagatat accatgggtc atcaccacca ccatcacggt ggcgctagca aaggtgaaga
     1441 gctgtttacg ggtgtagtac cgatcttagt ggaattagac ggcgacgtga acggtcacaa
     1501 atttagcgtg cgcggcgaag gcgaaggtga cgctaccaat ggtaaattga ccctgaagtt
     1561 tatttgcaca acaggcaaat tacccgttcc gtggcccacc ttagtgacca ccctgaccta
     1621 tggcgttcag tgcttcagtc gttacccaga tcatatgaaa caacacgatt ttttcaaatc
     1681 agccatgcct gaaggatatg ttcaagagcg tacaatcagc ttcaaggacg atggcaccta
     1741 taaaacgcgt gcggaagtga aatttgaagg cgacacatta gtaaaccgta tcgaactgaa
     1801 aggtatcgac ttcaaagaag acggcaacat tttaggccat aagctggaat ataactttaa
     1861 ttctcataac gtgtatatta cggccgataa acagaaaaac ggtatcaagg caaatttcaa
     1921 aattcgccat aacgtggaag acggcagcgt tcaattagcg gatcattatc aacaaaacac
     1981 gccgattggt gacgggcctg tactgttacc tgacaaccac tacctgagca cccagtcagc
     2041 actgagcaaa gatccgaacg aaaaacgcga tcacatggtt ctgttagaat tcgtgaccgc
     2101 tgcaggcatt actcacggaa tggacgagct gtacaaggat gaggaagagg acgattaata
     2161 actcgagtct ggtaaagaaa ccgctgctgc gaaatttgaa cgccagcaca tggactcgtc
     2221 tactagcgca gcttaattaa cctaggctgc tgccaccgct gagcaataac tagcataacc
     2281 ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg
     2341 attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     2401 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     2461 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     2521 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     2581 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     2641 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     2701 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     2761 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttc tggcggcacg
     2821 atggcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     2881 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     2941 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     3001 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     3061 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     3121 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     3181 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     3241 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     3301 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     3361 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     3421 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     3481 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     3541 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     3601 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     3661 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     3721 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     3781 tcatactctt cctttttcaa tcatgattga agcatttatc agggttattg tctcatgagc
     3841 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gtcatgacca aaatccctta
     3901 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     3961 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     4021 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     4081 cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa
     4141 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     4201 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     4261 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     4321 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     4381 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     4441 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     4501 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     4561 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     4621 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     4681 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg
     4741 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca
     4801 atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta cgtgactggg
     4861 tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     4921 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     4981 tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt
     5041 gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa
     5101 gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt tcctgtttgg
     5161 tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata ccgatgaaac
     5221 gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta ctggaacgtt
     5281 gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc actcagggtc
     5341 aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag cagcatcctg
     5401 cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc agactttacg
     5461 aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc
     5521 agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta aggcaacccc
     5581 gccagcctag ccgggtcctc aacgacagga gcacgatcat gctagtcatg ccccgcgccc
     5641 accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct
     5701 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     5761 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     5821 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     5881 accgcctggc cctgagagag tt
//
