LOCUS       sequence-338563-        6275 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a fusion protein of Gluc and iLID, which dimerizes with a 
            sspB-tagged protein when the bioluminescent reaction is initiated, 
            in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6275)
  AUTHORS   Adir O, Albalak MR, Abel R, Weiss LE, Chen G, Gruber A, Staufer O, 
            Kurman Y, Kaminer I, Shklover J, Shainsky-Roitman J, Platzman I, 
            Gepstein L, Shechtman Y, Horwitz BA, Schroeder A
  TITLE     Synthetic cells with self-activating optogenetic proteins 
            communicate with natural cells.
  JOURNAL   Nat Commun. 2022 Apr 28;13(1):2328. doi: 10.1038/s41467-022-29871-8.
  PUBMED    35484097
REFERENCE   2  (bases 1 to 6275)
  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: "Nat 
            Commun."; date: "2022-04-28"; volume: "13(1):2328. doi"; pages: " 
            10.1038/s41467-022-29871-8"
COMMENT     Sequence Label: pET28a-his-Gluc-iLID
FEATURES             Location/Qualifiers
     source          1..6275
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     965..984
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1004..1081)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1287..1306
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1390..1409
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1390..1408
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1409..1433
                     /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             1448..1470
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1481..1498
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2048..2485
                     /codon_start=1
                     /product="improved light-inducible dimer when used with 
                     SspB nano or SspB micro (Guntas et al., 2015)"
                     /label=iLID
                     /translation="LATTLERIEKNFVITDPRLPDNPIIFASDSFLQLTEYSREEILGR
                     NCRFLQGPETDRATVRKIRDAIDNQTEVTVQLINYTKSGKKFWNVFHLQPMRDYKGDVQ
                     YFIGVQLDGTERLHGAAEREAVMLIKKTAFQIAEAANDENYF"
     CDS             2529..2546
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2599..2617)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2613..2660
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2697..3152
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2784..2803)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2994..3015
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3244..4059)
                     /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(3244..4059)
                     /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     3967..3986
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4051..4070
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4181..4769
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4670..4689
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4923..4940
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4955..5094
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5020..5039)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5180..5202
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5196..5387)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6196..6275,1..1003))
                     /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 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
       61 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
      121 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
      181 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
      241 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
      301 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
      361 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
      421 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
      481 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
      541 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
      601 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
      661 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
      721 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
      781 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
      841 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
      901 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
      961 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     1021 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     1081 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     1141 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     1201 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     1261 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     1321 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
     1381 ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag
     1441 aaataatttt gtttaacttt aagaaggaga tatacatatg catcaccatc accatcacaa
     1501 accaacagaa aataacgaag attttaacat cgtcgcggta gcctcaaatt ttgcaaccac
     1561 cgacttagac gcggatcgtg ggaaattacc aggtaagaaa ttgccgttgg aagtacttaa
     1621 agaactggaa gccaacgcac gcaaggctgg atgcactcgt gggtgtttga tctgtctgtc
     1681 gcatattaag tgtacgccaa agatgaaaaa gttcatccca gggcgttgtc atacttacga
     1741 aggagacaaa gagtcagcac agggcggaat cggggaagct atcgtagata tccctgagat
     1801 cccaggcttc aaagacttgg aaccgctgga acagtttatt gctcaagttg acttgtgcgt
     1861 cgactgtacg actgggtgcc ttaaagggct ggcgaatgtc caatgtagtg atctgcttaa
     1921 gaagtggctt cctcagcgtt gcgctacatt cgcaagcaaa attcaggggc aagtggataa
     1981 gatcaaaggc gcgggtggtg acggtggtgg tggttcaggt ggtggtggtt caggatccgg
     2041 ggagtttctg gcaaccacac tggaacggat cgagaaaaat ttcgtgatta ctgatccgag
     2101 actgcctgac aacccaatca tttttgcgag cgattccttc ctgcagctga cagaatattc
     2161 tcgggaagag atcctggggc gcaattgccg ttttctgcag ggacccgaga cagaccgtgc
     2221 cactgttcgg aaaatcagag atgctattga caaccagact gaagtgaccg ttcagctgat
     2281 caattatacc aagagcggca agaagttctg gaacgtgttc cacctgcagc cgatgcgcga
     2341 ttataagggc gacgtccagt acttcattgg cgtgcagctg gatggcaccg aacgtcttca
     2401 tggcgccgct gagcgtgagg cggtcatgct gatcaaaaag acagcctttc agattgctga
     2461 ggcagcgaac gacgaaaatt acttttaaga attcgagctc cgtcgacaag cttgcggccg
     2521 cactcgagca ccaccaccac caccactgag atccggctgc taacaaagcc cgaaaggaag
     2581 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     2641 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg aatgggacgc
     2701 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2761 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2821 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2881 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2941 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3001 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     3061 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3121 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     3181 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa
     3241 ttcttagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc
     3301 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
     3361 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
     3421 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
     3481 gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg
     3541 ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga
     3601 ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat
     3661 cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
     3721 atattcttct aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc
     3781 atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca
     3841 gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
     3901 aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc
     3961 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
     4021 cggcctagag caagacgttt cccgttgaat atggctcata acaccccttg tattactgtt
     4081 tatgtaagca gacagtttta ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4141 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4201 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4261 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4321 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4381 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4441 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4501 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4561 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4621 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4681 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4741 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4801 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4861 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     4921 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     4981 atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc
     5041 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
     5101 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     5161 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     5221 aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg
     5281 tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt
     5341 ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg
     5401 taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg
     5461 atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg
     5521 gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt
     5581 aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac
     5641 ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag
     5701 accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc
     5761 tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc
     5821 ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc gataatggcc
     5881 tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc
     5941 aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc
     6001 tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag
     6061 acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg
     6121 ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca
     6181 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     6241 taatgaatcg gccaacgcgc ggggagaggc ggttt
//
