LOCUS       sequence-318813-        7738 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of N. tabacum Rubisco small subunit (rbcS-S1), Rubisco 
            chaperone protiens rbcX, raf1 and A. thaliana Rubisco chaperone 
            proteins raf2, and bsd2 in E. coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7738)
  AUTHORS   Lin MT, Stone WD, Chaudhari V, Hanson MR
  TITLE     Small subunits can determine enzyme kinetics of tobacco Rubisco 
            expressed in Escherichia coli.
  JOURNAL   Nat Plants. 2020 Oct;6(10):1289-1299. doi: 
            10.1038/s41477-020-00761-5. Epub 2020 Sep 14.
  PUBMED    32929197
REFERENCE   2  (bases 1 to 7738)
  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: 
            "10.1038/s41477-020-00761-5"; journalName: "Nat Plants"; date: 
            "2020-10"; volume: "6"; issue: "10"; pages: "1289-1299"
COMMENT     Sequence Label: pCDF-NtXR1AtR2B2NtS1
FEATURES             Location/Qualifiers
     source          1..7738
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      286..1024
                     /direction=RIGHT
                     /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."
     promoter        1072..1163
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1164..1955
                     /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"
     promoter        2069..2353
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     2276..2295
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     RBS             2380..2402
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     terminator      complement(2884..2931)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2983..3027)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     terminator      complement(3187..3234)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3302..3331)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="ASMTGGQQMG"
     RBS             3588..3610
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    3625..3649
                     /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)."
     promoter        complement(3650..3668)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      complement(3835..3882)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3950..3979)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="ASMTGGQQMG"
     RBS             4506..4528
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4543..4567
                     /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)."
     promoter        complement(4568..4586)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5939..5963
                     /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)."
     promoter        complement(5964..5982)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6565..6589
                     /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)."
     promoter        complement(6590..6608)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        6654..6731
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(6732..7738,1..76)
                     /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     complement(6751..6770)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
       61 ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat taggcaccgg
      121 gatctcgacc gatgcccttg agagccttca acccagtcag ctccttccgg tgggcgcggg
      181 gcatgactaa catgagaatt acaacttata tcgtatgggg ctgacttcag gtgctacatt
      241 tgaagagata aattgcactg aaatctagag cggttcagta gaaaagatca aaggatcttc
      301 ttgagatcct ttttttctgc gcgtaatctt ttgccctgta aacgaaaaaa ccacctgggg
      361 aggtggtttg atcgaaggtt aagtcagttg gggaactgct taaccgtggt aactggcttt
      421 cgcagagcac agcaaccaaa tctgtccttc cagtgtagcc ggactttggc gcacacttca
      481 agagcaaccg cgtgtttagc taaacaaatc ctctgcgaac tcccagttac caatggctgc
      541 tgccagtggc gttttaccgt gcttttccgg gttggactca agtgaacagt taccggataa
      601 ggcgcagcag tcgggctgaa cggggagttc ttgcttacag cccagcttgg agcgaacgac
      661 ctacaccgag ccgagatacc agtgtgtgag ctatgagaaa gcgccacact tcccgtaagg
      721 gagaaaggcg gaacaggtat ccggtaaacg gcagggtcgg aacaggagag cgcaagaggg
      781 agcgacccgc cggaaacggt ggggatcttt aagtcctgtc gggtttcgcc cgtactgtca
      841 gattcatggt tgagcctcac ggctcccaca gatgcaccgg aaaagcgtct gtttatgtga
      901 actctggcag gagggcggag cctatggaaa aacgccaccg gcgcggccct gctgttttgc
      961 ctcacatgtt agtcccctgc ttatccacgg aatctgtggg taactttgta tgtgtccgca
     1021 gcgcccgccg cagtctcacg cccggagcgt agcgaccgag tgagctagct atttgtttat
     1081 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     1141 aataatattg aaaaaggaag agtatgaggg aagcggtgat cgccgaagta tcgactcaac
     1201 tatcagaggt agttggcgtc atcgagcgcc atctcgaacc gacgttgctg gccgtacatt
     1261 tgtacggctc cgcagtggat ggcggcctga agccacacag tgatattgat ttgctggtta
     1321 cggtgaccgt aaggcttgat gaaacaacgc ggcgagcttt gatcaacgac cttttggaaa
     1381 cttcggcttc ccctggagag agcgagattc tccgcgctgt agaagtcacc attgttgtgc
     1441 acgacgacat cattccgtgg cgttatccag ctaagcgcga actgcaattt ggagaatggc
     1501 agcgcaatga cattcttgca ggtatcttcg agccagccac gatcgacatt gatctggcta
     1561 tcttgctgac aaaagcaaga gaacatagcg ttgccttggt aggtccagcg gcggaggaac
     1621 tctttgatcc ggttcctgaa caggatctat ttgaggcgct aaatgaaacc ttaacgctat
     1681 ggaactcgcc gcccgactgg gctggcgatg agcgaaatgt agtgcttacg ttgtcccgca
     1741 tttggtacag cgcagtaacc ggcaaaatcg cgccgaagga tgtcgctgcc gactgggcaa
     1801 tggagcgcct gccggcccag tatcagcccg tcatacttga agctagacag gcttatcttg
     1861 gacaagaaga agatcgcttg gcctcgcgcg cagatcagtt ggaagaattt gtccactacg
     1921 tgaaaggcga gatcaccaag gtagtcggca aataatgtct aacaattcgt tcaagccgag
     1981 gggccgcaag atccggccac gatgacccgg tcgtcggttc agggcagggt cgttaaatag
     2041 ccgcttatgt ctattgctgg tttaccggaa gaaaccaatt gtccatattg catcagacat
     2101 tgccgtcact gcgtctttta ctggctcttc tcgctaacca aaccggtaac cccgcttatt
     2161 aaaagcattc tgtaacaaag cgggaccaaa gccatgacaa aaacgcgtaa caaaagtgtc
     2221 tataatcacg gcagaaaagt ccacattgat tatttgcacg gcgtcacact ttgctatgcc
     2281 atagcatttt tatccataag attagcggat cctacctgac gctttttatc gcaactctct
     2341 actgtttctc catacccgtt tttttgggct agaaataatt ttgtttaact ttaagaagga
     2401 gatatacata cccatgcagg tgtggccacc aattaacaag aagaagtacg agactctctc
     2461 ataccttcct gatttgagcc aggagcaatt gcttagtgaa gttgagtacc ttttgaaaaa
     2521 tggatgggtt ccttgcttgg aattcgagac tgagcacgga tttgtctatc gtgaaaacaa
     2581 caagtcacca ggatactatg atggcagata ctggaccatg tggaagctac ctatgttcgg
     2641 atgcactgat gccacccaag tgttggctga ggtagaagag gcgaagaagg catacccaca
     2701 ggcctggatc cgtatcattg gattcgacaa cgtgcgtcaa gtgcagtgca tcagtttcat
     2761 tgcctacaag cctgaaggat attaagtcgc aaaaaacccc gcttcggcgg ggttttttcg
     2821 ctccggttta ttgactaccg gaagcagtgt gaccgtgtgc ttctcaaatg cctgaggttt
     2881 cagcaaaaaa cccctcaaga cccgtttaga ggccccaagg ggttatgcta gttattgctc
     2941 agcggtggca gcagcctagg ttaattaagc tgcgctagta gacgagtcca tgtgctggcg
     3001 ttcaaatttc gcagcagcgg tttctttacc agactcgagg gtaccgacgt cagcgatcgc
     3061 gtggccggcc gatatccaat tccgcatgcg ctagcacctc cactacaatt gaccgctggt
     3121 accgcttggt tatgccggta ctgccgggcc tcttgcggga tatccggata tagttcctcc
     3181 tttcagcaaa aaacccctca agacccgttt agaggcccca aggggttatg ctagttattg
     3241 ctcagcggtg gcagcagcca actcagcttc ctttcgggct ttgttagcag ccggatctgc
     3301 gacccatttg ctgtccacca gtcatgctag cctactcatc gaaggtgctc aagaacccac
     3361 caatgaaacc agctccattg cagtccccac acaacacctc ctttttacct ctgcaaagcc
     3421 aacagagtgc accagcttta aactgaccat tgaaatgatc aatcaaattc acaccacctc
     3481 ctttgcattg agaacatgcc acacaacctt ccccttcaca atttgcacaa actaaactgt
     3541 ttggtttagt gccctgagga ttattgtttg cggccatcat atgtatatct ccttcttaaa
     3601 gttaaacaaa attatttcta gaggggaatt gttatccgct cacaattccc ctatagtgag
     3661 tcgtattaat ttcgcgggat cgagatctcg atcctctacg ccggacgcat cgtggccggc
     3721 atcaccggcg ccacaggtgc ggttgctggc gcctatatcg ccgacactag tgcttggtta
     3781 tgccggtact gccgggcctc ttgcgggata tccggatata gttcctcctt tcagcaaaaa
     3841 acccctcaag acccgtttag aggccccaag gggttatgct agttattgct cagcggtggc
     3901 agcagccaac tcagcttcct ttcgggcttt gttagcagcc ggatccgcga cccatttgct
     3961 gtccaccagt catgctagct cacgcccaag ctcttttcct aggggaaaga tcagaagtct
     4021 tgatatcatc tattttagcc gccattatga aatcattcat gctcaaccct ccgatagaag
     4081 aggtaaatag ttcagctcga acttgggtag gactttccaa gtgaagagaa gggtaatgac
     4141 cagaagcttc agcaacctta tggatcctgt ttatgagttc aacaccgcac ccaaaatccc
     4201 tcactttcca catacacctt attttcagac cacccgcttc attatccact atactccatc
     4261 ccaaaacctt gtggaggaga gttctggcat catcgccgga caaaggaggc tgtgaagacg
     4321 aaacagggga acactgaagc tgggagagag acaaaacaga tgcgtttggt atcaaaatct
     4381 tgtgctcagt gctttggcat cccaacactt tatctccgaa ttgactcgct atctcctccg
     4441 ggtaagggtc acgagctccg aagtcaccaa gaaaatcctg cgccagatta gacatcatat
     4501 gtatatctcc ttcttaaagt taaacaaaat tatttctaga ggggaattgt tatccgctca
     4561 caattcccct atagtgagtc gtattaattt cgcgggatcg agatctcgat cctctacgcc
     4621 ggacgcatcg tggccggcat caccggcgcc acaggtgcgg ttgctggcgc ctatatcgcc
     4681 gacctgcagt ctagaaagct aagcttcttg gtctttaatc ccaatcttct tcacctaatt
     4741 gttcttcatc ttcccacctt ggtggcctaa ctactataag cacaattcct aagctctgtt
     4801 caacccctct tttcttcaat gttgaccctc tttccacttt caatccctcc tcattgctcc
     4861 cgtctccacc atcaaccacc aagtaaaacc cgtcctccga cgccacttct gtccttcccc
     4921 tatccgccac taccaaaacc ggctcttgct tgtacttact cttctccctc cacggcaaaa
     4981 agttcccgct cttgaacgac actgcaactc cacctctctc taaccccgcg ataggttgcc
     5041 atccgggcaa aacaacccat ctcctccaat ccttctccgc ctccactata ccaaaatccc
     5101 ctacgccacc acattcccac ggcgccgcct ccacctccac gtcccttccc tccgccttac
     5161 aaacaggcaa aacgactaca atcgtagact cagctacctc ccctaacttc attctcacca
     5221 aaggcactgt caccccatca tcaagtacac tctccttagc aacctcacgt ccttctagct
     5281 ccatcacaag caaattcctc gcactttcag actcaacaac ttctaatgcc ttctccattg
     5341 acgacctcct caaatcctca gaagctgccg cagcatattc ttgagctaat ctaaagtaca
     5401 taaaagctaa acaatcacca ggagaatcac cattaaactt gtcccaacca taatccacac
     5461 gtctacgcgg ataatcttta attgctctag ctaaatcttg agcttgtttt gcatcaaacc
     5521 cattcttcac taaaaatgtt gctgcatcag ttcgttgcct agcactaagc aaacgaattt
     5581 catataataa ttcagcacct ccagattcaa aatacgacaa cgtttcttca tctaaagcag
     5641 cagactcaac aagagtatca cgaacttggg cagcaacaac caaccgattc tgttcaactc
     5701 cagtaaggcc agtgatttcc tcaagagttg aaggagtaaa cccttcactt gttaaataag
     5761 gtatcaaagg agcatattca taccaaagac caaggcgatt agatagaact tcgagctttg
     5821 catttgtgtc aagattgcgg aatttgggtg ggagaggtgg aggaggaggt ctaaaaggct
     5881 gataaagttg ttgcatatgt atatctcctt cttatactta actaatatac taagatgggg
     5941 aattgttatc cgctcacaat tcccctatag tgagtcgtat taatttcgat tatgcggccg
     6001 tgtacaatac gattactttc tgttcgactt aagcattatg cggccgcact gggcgcgcct
     6061 cacccaactc tctccgagtt cgtgtcatct gatggccaaa tcacagtttc catgagccgc
     6121 tcacgcatca cttccacatt ctgatcagat attcggttgt agatctcctc gtgatcacat
     6181 ttttttatcc atttcgcata cagactgagc cgtgttacca tcactttttc agcgagttct
     6241 tgcttctcct tagcaaggtt gcgaataaaa cgcttcccat ccctaagctt atttactgta
     6301 acataatcat tgaaccattg atattgtgga gagtccattt cattaagctg gttcaaaata
     6361 gtcctcactg ctttaaatgt gaagtagtta agaatttgcg ttttgatatc atcaaagcca
     6421 tattcatact gtccacctaa atcactgaca accacaagat tcctcctact ctttttctcc
     6481 ttctttaaat tcttgtaatt acccttaaat gcttttgcca tggtatatct ccttattaaa
     6541 gttaaacaaa attatttcta caggggaatt gttatccgct cacaattccc ctatagtgag
     6601 tcgtattaat ttcctaatgc aggagtcgca taagggagag cgtcgagatc ccggacacca
     6661 tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
     6721 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
     6781 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
     6841 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
     6901 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
     6961 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
     7021 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
     7081 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
     7141 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca
     7201 acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat
     7261 tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc
     7321 gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg
     7381 aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca
     7441 tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca
     7501 ttaccgagtc cgggctgcgc gttggtgcgg acatctcggt agtgggatac gacgataccg
     7561 aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg
     7621 ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc
     7681 agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaac
//
