LOCUS       sequence-346285-        4898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NanoLuc enzyme linked with SynZip2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4898)
  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 4898)
  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: NanoLuc-SZ2
FEATURES             Location/Qualifiers
     source          1..4898
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..78)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     284..303
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     387..406
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        387..405
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    406..430
                     /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             445..467
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             478..990
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             1003..1023
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1180..1197
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1250..1268)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1264..1311
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1348..1803
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1435..1454)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1645..1666
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1918..2733)
                     /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(1918..2733)
                     /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     2641..2660
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2725..2744
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2886..3474
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3375..3394
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3628..3645
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3816..4898)
                     /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     4860..4879
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
       61 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
      121 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
      181 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
      241 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
      301 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
      361 ggatcgagat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
      421 taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat acatatgatg
      481 gtcttcacac tcgaagattt cgttggggac tggcgacaga cagccggcta caacctggac
      541 caagtccttg aacagggagg tgtgtccagt ttgtttcaga atctcggggt gtccgtaact
      601 ccgatccaaa ggattgtcct gagcggtgaa aatgggctga agatcgacat ccatgtcatc
      661 atcccgtatg aaggtctgag cggcgaccaa atgggccaga tcgaaaaaat ttttaaggtg
      721 gtgtaccctg tggatgatca tcactttaag gtgatcctgc actatggcac actggtaatc
      781 gacggggtta cgccgaacat gatcgactat ttcggacggc cgtatgaagg catcgccgtg
      841 ttcgacggca aaaagatcac tgtaacaggg accctgtgga acggcaacaa aattatcgac
      901 gagcgcctga tcaaccccga cggctccctg ctgttccgag taaccatcaa cggagtgacc
      961 ggctggcggc tgtgcgaacg cattctggcg ggcggcggat ctgagaattt atatttccag
     1021 ggcgctcgta atgcatacct gcgtaaaaaa atcgcgcgtt taaaaaaaga caacttgcaa
     1081 cttgagcgcg acgaacaaaa tttggagaaa ataatcgcca accttcggga cgagatcgcg
     1141 cgtctggaaa acgaggtggc ttcgcatgag caactcgagc accaccacca ccaccactga
     1201 gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa
     1261 taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagga
     1321 ggaactatat ccggattggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     1381 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     1441 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     1501 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     1561 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     1621 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     1681 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     1741 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     1801 tttaggtggc acttttcggg gtacccaggt ggcacttttc ggggaaatgt gcgcggaacc
     1861 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgaa ttaattctta
     1921 gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc
     1981 atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag
     2041 gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat
     2101 taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga
     2161 atccggtgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc
     2221 attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc
     2281 ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg
     2341 caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc
     2401 ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc
     2461 aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag
     2521 tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa
     2581 ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt
     2641 atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta atcgcggcct
     2701 agagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta
     2761 agcagacagt tttattgttc atgaccaaaa tcccttaacg tgacctaggt catgaccaaa
     2821 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     2881 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     2941 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     3001 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     3061 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     3121 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     3181 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     3241 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     3301 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     3361 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     3421 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     3481 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     3541 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     3601 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     3661 cataatccat gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc
     3721 accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct
     3781 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     3841 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3901 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     3961 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
     4021 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
     4081 tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
     4141 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
     4201 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
     4261 atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc
     4321 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
     4381 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
     4441 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
     4501 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
     4561 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
     4621 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
     4681 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
     4741 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
     4801 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
     4861 gcatactctg cgacatcgta taacgttact ggtttcac
//
