LOCUS       sequence-346286-        5741 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NanoLuc enzyme fused to LAF-1 RGG domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5741)
  AUTHORS   Guan M, Garabedian MV, Leutenegger M, Schuster BS, Good MC, Hammer 
            DA
  TITLE     Incorporation and Assembly of a Light-Emitting Enzymatic Reaction 
            into Model Protein Condensates.
  JOURNAL   Biochemistry. 2021 Oct 14. doi: 10.1021/acs.biochem.1c00373.
  PUBMED    34648259
REFERENCE   2  (bases 1 to 5741)
  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: 
            "Biochemistry. 2021 Oct 14. doi: 10.1021/acs.biochem.1c00373."
COMMENT     Sequence Label: RGG-NanoLuc-RGG
FEATURES             Location/Qualifiers
     source          1..5741
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             181..693
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             1219..1236
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1289..1307)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1303..1350
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1387..1842
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1474..1493)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1684..1705
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1957..2772)
                     /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(1957..2772)
                     /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"
     primer_bind     2680..2699
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2764..2783
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2925..3513
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3414..3433
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3667..3684
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3855..4937)
                     /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     4899..4918
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4938..5015)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5221..5240
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5324..5343
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5324..5342
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5343..5367
                     /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             5382..5404
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 ggctataaca ataaccgtgg tggcggtggc ggtggctaca atcgccagga tcgtggtgac
       61 ggtggcagct ctaacttttc gcgcggtggc tataataacc gtgatgaagg cagcgacaac
      121 cgcggtagtg gccgttccta caataacgat cgtcgcgaca atggtggcga tggcgaattc
      181 atggtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg
      241 gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta
      301 actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc
      361 atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag
      421 gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta
      481 atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc
      541 gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc
      601 gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg
      661 accggctggc ggctgtgcga acgcattctg gcgacattcg ggaagcttat ggaatccaat
      721 caatccaata acggtggctc gggtaatgcc gctctgaatc gtggtggccg ctacgtgccg
      781 ccgcatctgc gtggtggcga tggtggtgca gcagcagctg catctgctgg cggtgatgac
      841 cgtcgcggcg gtgcgggcgg tggcggttat cgtcgcggcg gtggcaatag tggtggcggt
      901 ggcggtggcg gttatgatcg tggttacaat gacaaccgtg atgaccgcga taaccgtggc
      961 ggttccggcg gttacggtcg tgatcgtaat tatgaagacc gtggttacaa cggcggtggc
     1021 ggtggcggtg gcaatcgtgg ctataacaat aaccgtggtg gcggtggcgg tggctacaat
     1081 cgccaggatc gtggtgacgg tggcagctct aacttttcgc gcggtggcta taataaccgt
     1141 gatgaaggca gcgacaaccg cggtagtggc cgttcctaca ataacgatcg tcgcgacaat
     1201 ggtggcgatg gcctcgagca ccaccaccac caccactgag atccggctgc taacaaagcc
     1261 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
     1321 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg
     1381 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     1441 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     1501 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     1561 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     1621 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     1681 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     1741 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     1801 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     1861 tacccaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     1921 atacattcaa atatgtatcc gctcatgaat taattcttag aaaaactcat cgagcatcaa
     1981 atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa aaagccgttt
     2041 ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat cctggtatcg
     2101 gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct cgtcaaaaat
     2161 aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga atggcaaaag
     2221 tttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt catcaaaatc
     2281 actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgagac gaaatacgcg
     2341 atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca ggaacactgc
     2401 cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct ggaatgctgt
     2461 tttcccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga taaaatgctt
     2521 gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct catctgtaac
     2581 atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat cgggcttccc
     2641 atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc atttataccc
     2701 atataaatca gcatccatgt tggaatttaa tcgcggccta gagcaagacg tttcccgttg
     2761 aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt ttattgttca
     2821 tgaccaaaat cccttaacgt gacctaggtc atgaccaaaa tcccttaacg tgagttttcg
     2881 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     2941 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     3001 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     3061 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     3121 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     3181 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     3241 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     3301 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     3361 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     3421 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     3481 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     3541 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     3601 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     3661 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc ataatccatg ataaagaaga
     3721 cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag ctgactgggt
     3781 tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat
     3841 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     3901 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt
     3961 cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt
     4021 tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt
     4081 aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc
     4141 gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg
     4201 ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga
     4261 aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga
     4321 gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc
     4381 gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta
     4441 ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat
     4501 aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga
     4561 tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag
     4621 gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg
     4681 cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca
     4741 acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa
     4801 ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc
     4861 tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat
     4921 aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc
     4981 ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg
     5041 cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc
     5101 aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac
     5161 catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc
     5221 ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac
     5281 gatgcgtccg gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata
     5341 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
     5401 gagatataca tatggaatcc aatcaatcca ataacggtgg ctcgggtaat gccgctctga
     5461 atcgtggtgg ccgctacgtg ccgccgcatc tgcgtggtgg cgatggtggt gcagcagcag
     5521 ctgcatctgc tggcggtgat gaccgtcgcg gcggtgcggg cggtggcggt tatcgtcgcg
     5581 gcggtggcaa tagtggtggc ggtggcggtg gcggttatga tcgtggttac aatgacaacc
     5641 gtgatgaccg cgataaccgt ggcggttccg gcggttacgg tcgtgatcgt aattatgaag
     5701 accgtggtta caacggcggt ggcggtggcg gtggcaatcg t
//
