LOCUS       sequence-474338-        6333 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Anthoceros agrestis chaperones: Raf1,Raf2,RbcX2,BSD2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6333)
  AUTHORS   Oh ZG, Robison TA, Loh DH, Ang WSL, Ng JZY, Li FW, Gunn LH
  TITLE     Unique biogenesis and kinetics of hornwort Rubiscos revealed by 
            synthetic biology systems.
  JOURNAL   Mol Plant. 2024 Dec 2;17(12):1833-1849. doi: 
            10.1016/j.molp.2024.10.013. Epub 2024 Nov 2.
  PUBMED    39491367
REFERENCE   2  (bases 1 to 6333)
  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: 
            "10.1016/j.molp.2024.10.013"; journalName: "Mol Plant"; date: 
            "2024-12-2- 2"; volume: "17"; issue: "12"; pages: "1833-1849"
COMMENT     Sequence Label: pCDF_AaRaf1/Raf2/RbcX2/BSD2
FEATURES             Location/Qualifiers
     source          1..6333
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(700..791)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(839..1577)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     CDS             complement(1787..2869)
                     /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     2831..2850
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2870..2947)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        2989..3007
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3008..3032
                     /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)."
     terminator      4293..4340
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        4367..4385
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4386..4410
                     /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             4425..4447
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     terminator      5009..5056
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        5105..5123
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5124..5148
                     /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             5163..5185
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     terminator      5681..5728
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        5773..5791
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5792..5816
                     /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             5831..5853
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     terminator      6169..6216
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(join(6241..6333,1..699))
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
ORIGIN
        1 tccaagataa gcctgtctag cttcaagtat gacgggctga tactgggccg gcaggcgctc
       61 cattgcccag tcggcagcga catccttcgg cgcgattttg ccggttactg cgctgtacca
      121 aatgcgggac aacgtaagca ctacatttcg ctcatcgcca gcccagtcgg gcggcgagtt
      181 ccatagcgtt aaggtttcat ttagcgcctc aaatagatcc tgttcaggaa ccggatcaaa
      241 gagttcctcc gccgctggac ctaccaaggc aacgctatgt tctcttgctt ttgtcagcaa
      301 gatagccaga tcaatgtcga tcgtggctgg ctcgaagata cctgcaagaa tgtcattgcg
      361 ctgccattct ccaaattgca gttcgcgctt agctggataa cgccacggaa tgatgtcgtc
      421 gtgcacaaca atggtgactt ctacagcgcg gagaatctcg ctctctccag gggaagccga
      481 agtttccaaa aggtcgttga tcaaagctcg ccgcgttgtt tcatcaagcc ttacggtcac
      541 cgtaaccagc aaatcaatat cactgtgtgg cttcaggccg ccatccactg cggagccgta
      601 caaatgtacg gccagcaacg tcggttcgag atggcgctcg atgacgccaa ctacctctga
      661 tagttgagtc gatacttcgg cgatcaccgc ttccctcata ctcttccttt ttcaatatta
      721 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
      781 aaataaacaa atagctagct cactcggtcg ctacgctccg ggcgtgagac tgcggcgggc
      841 gctgcggaca catacaaagt tacccacaga ttccgtggat aagcagggga ctaacatgtg
      901 aggcaaaaca gcagggccgc gccggtggcg tttttccata ggctccgccc tcctgccaga
      961 gttcacataa acagacgctt ttccggtgca tctgtgggag ccgtgaggct caaccatgaa
     1021 tctgacagta cgggcgaaac ccgacaggac ttaaagatcc ccaccgtttc cggcgggtcg
     1081 ctccctcttg cgctctcctg ttccgaccct gccgtttacc ggatacctgt tccgcctttc
     1141 tcccttacgg gaagtgtggc gctttctcat agctcacaca ctggtatctc ggctcggtgt
     1201 aggtcgttcg ctccaagctg ggctgtaagc aagaactccc cgttcagccc gactgctgcg
     1261 ccttatccgg taactgttca cttgagtcca acccggaaaa gcacggtaaa acgccactgg
     1321 cagcagccat tggtaactgg gagttcgcag aggatttgtt tagctaaaca cgcggttgct
     1381 cttgaagtgt gcgccaaagt ccggctacac tggaaggaca gatttggttg ctgtgctctg
     1441 cgaaagccag ttaccacggt taagcagttc cccaactgac ttaaccttcg atcaaaccac
     1501 ctccccaggt ggttttttcg tttacagggc aaaagattac gcgcagaaaa aaaggatctc
     1561 aagaagatcc tttgatcttt tctactgaac cgctctagat ttcagtgcaa tttatctctt
     1621 caaatgtagc acctgaagtc agccccatac gatataagtt gtaattctca tgttagtcat
     1681 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga
     1741 tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     1801 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     1861 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
     1921 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     1981 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     2041 cggtatcgtc gtatcccact accgagatgt ccgcaccaac gcgcagcccg gactcggtaa
     2101 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     2161 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     2221 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     2281 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     2341 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     2401 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     2461 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     2521 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     2581 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     2641 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     2701 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     2761 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     2821 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     2881 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     2941 tggtgtccgg gtctcaggag cgggatctcg acgcctgcat taggaaatta atacgactca
     3001 ctatagggga attgtgagcg gataacaatt cccctgtaga aataattttg tttaacttta
     3061 ataaggagat ataccatggg ggggagggac gtttaccagc ctttccggcc cccggtcacc
     3121 aacctgcctg ctccgggtac cgtgtccccg gaggagcagc tgcagatcct gagggagagg
     3181 cgggggctat ggcacgagta cgcgctggtc atccccttcc tcacccgagc aggctacact
     3241 ccggcgcaga tcgatgaggc aaccggggtg accggggtgg agcagaacat gatcgtgggg
     3301 gcgtcccagg tgcggctgtc gctcatagcc gggggaatgg gagcggaggt gctgaatttc
     3361 ttcgatgtgg gaggggcaga cctgctgtac gagctcagaa ttctgtccac ggatcagagg
     3421 aaggcttcgg cggagttgat cgtggagaaa aagatggacc ctaaagaggc caaggagctg
     3481 gctcgggctg tgaaggacta cgataggagg aagagggagg aggggtgggg atccttcaac
     3541 aacggcaggc cgggggacta tcttgccttc gcttgttacc gccagagcaa ggaggtgaga
     3601 ttcgatgctg agaaagagat tctgatcaac aaggggctca gctatgccgc ctctgactct
     3661 gcacggacca agttgaagga agccctaggg ttggaagcag ctccggtgtc acccgagcgg
     3721 gaggaggccg ttagggagga ggccgttagg tctaggttac aggtggtgag gcttcctagt
     3781 aagcaggaca cggaaaactc tcccctgcct gtcatactgc ctgtagctac tgacctggcg
     3841 actgtgaaaa aagctcctgc ggttggagct tttggtgatg gtccttttaa cgtttacggt
     3901 gcacagccta gatggaagac atggctagct cttccaggat ggtcctctgt cgtgacgtcc
     3961 cgggctccca ttgctgtggt gttccctgat gcaactatct tagatgtggg gtcaactccg
     4021 ggaactgttg caaggcaggg cagaaacgag cctattttgt tggtggtgga caagggaaac
     4081 gtcgatccgg aggaagatgc ttactttctg gtggaggaaa accagcgctt ggctttgcgg
     4141 ttaggttcga aaccagctgc cacaacccca aagaactcaa agacattcgg gaaagttgta
     4201 gttgtattga gaccccccct ttctgaggac gaggatgcag tttgggaatg atcgaccagg
     4261 cattactaga gtcacactcc gctgagcaat aactagcata accccttggg gcctctaaac
     4321 gggtcttgag gggttttttg aatgcgagat ctcgatcccg cgaaattaat acgactcact
     4381 ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag
     4441 aaggagatat acatatggca caggccgtgt cgtcggatcc tttgggagag gttggagcca
     4501 gggatccaac atcctctgaa atcgagagca gattcggaga taaggtgttg ggaaatgcca
     4561 gcacggagca cgtgatcctc attccgccgc acctcctggg cctcgccgac cgcgccgtca
     4621 caccccttgc tccaggcacg cagcctctat ccgctgacgc ggccaaagct ctacttcgca
     4681 aggtagttgc ctggagactt atcgaccagg gaggccttcg catcgagtgc gagtggaagc
     4741 tcaagagttt tggtgctggc attgagctgt tcgaacgtat agctgaggtg gcggaggccg
     4801 ccggtcacca ccctgacctt catctcgaac aaggcaacag agccagagtc tgcttgtcgt
     4861 ctcactccgt cggaggcttg acggagaatg acttcattgt agctgccagg atagatcgcg
     4921 tgaagacatc ggatctcata ttcaagaaga acaagttctg ggcctgatcg accaggcatt
     4981 actagagtca cactccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     5041 cttgaggggt tttttgaggt cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg
     5101 aaattaatac gactcactat aggggaattg tgagcggata acaattcccc tctagaaata
     5161 attttgttta actttaagaa ggagatatac atatggtgcc ttcatgtggt caacaaggat
     5221 cacggagacc tcttgttgta tgtgagcttg gtgggaatta tgaggaaagt tttgcagatg
     5281 ttgacaagca ccttcttgat tatttcacgt tcaaagcggt gaagactgtt ttggcacagt
     5341 tgtcagagat gaaccctgag cagtattcct ggttttataa ctttgttgtg agtaatagac
     5401 ctcaagatag caaatacttt gtgcgcctgt tagtgaagga aaggcaagag cttggtgaga
     5461 gggtcatggt aactagacta catcttttca acgcatgggt gaagaagtac aaccatgcgc
     5521 agatgcacaa ggccatctca gagcagaacc tcgagattct gagggaacgt ctcatccaaa
     5581 cagtgaaatt gacatcagac ggggatgact tcgacaggcc acctttgagg ggccagtaat
     5641 cgaccaggca ttactagagt cacactccgc tgagcaataa ctagcataac cccttggggc
     5701 ctctaaacgg gtcttgaggg gttttttggc ttgcgtccgg cgtagaggat cgagatctcg
     5761 atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac aattcccctc
     5821 tagaaataat tttgtttaac tttaagaagg agatatacat atggcctctg cgcaggggga
     5881 tgagtctaag gcgacctcgc agtcgggagt gaagacaaag agcattcttt gcatgaactg
     5941 cacggggaac ggggctgttg tgtgctccca gtgtaaggga aaaggcttga attcggtgga
     6001 tcacttcaac ggccggtaca aagcaggcca aatttgctgg ctgtgcaggg gcaagaaaca
     6061 gatgctctgc ggagagtgca acggtgccgg attcattggt ggctttctga atacgcaaga
     6121 cgattagtcg accaggcatt actagagtca cactccgctg agcaataact agcataaccc
     6181 cttggggcct ctaaacgggt cttgaggggt tttttgggtc tccgcaagaa ttgttagaca
     6241 ttatttgccg actaccttgg tgatctcgcc tttcacgtag tggacaaatt cttccaactg
     6301 atctgcgcgc gaggccaagc gatcttcttc ttg
//
