LOCUS       sequence-318481-        9201 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of N. tabacum Rubisco large subunit (rbcL) and Rubisco 
            chaperone protiens cpn60-alpha, cpn60-beta, cpn20 in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9201)
  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 9201)
  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: pET-NtC60-alpha-C20NtLC60-beta
FEATURES             Location/Qualifiers
     source          1..9201
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(17..35)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        103..207
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(208..276,277..1068)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(426..445)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1239..1827
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1728..1747
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1970..1987
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2002..2141
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2067..2086)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2227..2249
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2243..2434)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        3113..3131
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3132..3156
                     /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             3171..3193
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4205..4216
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     RBS             4863..4885
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     promoter        5520..5538
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5539..5563
                     /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             5578..5600
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     promoter        7128..7146
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7147..7171
                     /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)."
     primer_bind     complement(8923..8941)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      8937..8984
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
       61 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
      121 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
      181 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
      241 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
      301 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
      361 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
      421 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
      481 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
      541 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
      601 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
      661 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
      721 ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta
      781 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
      841 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
      901 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
      961 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     1021 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     1081 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     1141 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     1201 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     1261 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     1321 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     1381 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     1441 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     1501 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     1561 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     1621 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     1681 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     1741 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     1801 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     1861 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     1921 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     1981 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     2041 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     2101 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     2161 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     2221 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     2281 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     2341 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     2401 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     2461 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     2521 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     2581 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     2641 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     2701 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     2761 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     2821 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     2881 gcacgatcat gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc
     2941 tgctggagat ggcggacgcg atggatatgt tctgccaagg gttggtttgc gcattcacag
     3001 ttctccgcaa gaattgattg gctccaattc ttggagtggt gaatccgtta gcgaggtgcc
     3061 gccggccacg atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga
     3121 ctcactatag gggaattgtg agcggataac aattcccctc tagcaataat tttgtttaac
     3181 tttaagaagg agatatacat atggctaaag aaattgcctt tgaccagaaa tctaggagtg
     3241 cccttcaggc tggtatcgat aagctcgctg atgctgtcgg tctcactctt ggtcccaggg
     3301 ggaggaatgt agtgttggat gaatatggca ccccaaaggt ggttaatgac ggagttacaa
     3361 ttgctcgtgc tatagagcta cctgatgcta tggaaaatgc tggcgctgcc cttatcaggg
     3421 aggttgcaag caaaactaac gattcagccg gtgatggaac cacgactgca tctgttcttg
     3481 ctcgcgaaat cattaaactt ggtctattga gtgttacatc tggtgcaaat ccagtgtctt
     3541 tgaagaaggg cattgacaag actgtactgg gtttgattga agagctagaa aagaaggcta
     3601 gacctgttaa aggtcgtgat gacatcaaag ctattgcttc gatctctgct ggaaatgatg
     3661 aaagcattgg caccatgatt gcggatgcaa ttgacaaagt cggtccagat ggtgtgttgt
     3721 ccatcgagtc atcctcctcc tttgagacaa ctgttcatgt tgaagaagga atggagattg
     3781 atagaggata tatttcccca caatttgtca ccaaccctga gaaattaatt gttgagtttg
     3841 agaatgctag agtcttagtt acggatcaga agatctcggc tatcaaggat attatccccc
     3901 tattggaaaa gacaactcag ttacgctctc ctctgctcat tattgctgag gatgtcaccg
     3961 gggaagctct ggctactctt gtcgtgaaca agctgcgggg tatactgaat gttgccgcca
     4021 tcaaagctcc tggatttggt gaaaggagga aggctcttct gcaagatatt gccattgtga
     4081 caggggctga gtaccaagca actgacctgg gcctgctcat tgagggtacc ccagttgaag
     4141 cacttggaat tgccagaaag gtgaccatta ccaaggattc aacaaccatc attgctgatg
     4201 cagcatcgaa ggacgagata caggctagga ttgctcagct caaaaaggag ttgtccgaga
     4261 cagactcggt ttatgactcc gagaaacttg ctgagagaat cgccaagttg tctgggggtg
     4321 ttgccgtcat caaggtcgga gctgctacag aggctgagct tgaggaccgc aagcttcgca
     4381 tcgaggatgc aaagaatgca acttttgctg caatcgaaga gggaatagta cctggtggtg
     4441 gtgctgcctt tgttcatcta tcaacttatg tccctgccat taaggacaag ctcgaagatg
     4501 cagatgaaaa gttgggagct gacatcattc aaagggcatt agtagcacca gcatctttga
     4561 tagcccaaaa tgctggaatt gaaggggaag tagtagtgga gaaggtaagg gaagctgaat
     4621 gggagatggg ttataacgcg atgacagaca aatacgagaa tcttgttgat gctggagtga
     4681 tcgatccagc caaggtgaca agatgtgcat tgcagaattc agcatcagtt gcaggaatgg
     4741 tacttacaac acaagccatt gtggtcgaga aggcaaagcc taaggcagct gcacctgctg
     4801 ctccacaagg gctctcagtg tgatacttgg cgcgcggata acaattgccc tctagaaata
     4861 attttgttta actttaagaa ggagatatac atatggctgc tactgttgta gcccccaaat
     4921 acacttctct taagcctttg ggtgatagag ttttggtgaa gattaagact gtagaggaaa
     4981 agaccgtagg tggtatccta cttccaacaa cagcacagtc gaaacctcaa ggaggtgagg
     5041 ttgttgctgt tggggagggt cgttcagttg ggaagaatca ggtggacatt agtgtgaaga
     5101 ctggcaccca agttgtttac tcaaaatatg cgggaacaga ggtggagttc aatggatcaa
     5161 atcacctcat cttgaaggag gatgacattg ttggtattct cgagacagat gaaatcaagg
     5221 atctgcagcc cttgaatgac agagttctga taaaggtcgc cgaggctgag gaaaaaactg
     5281 ctggaggctt attattaact gaagcatcca aggagaagcc ttccattggc acggttatag
     5341 ctgtaggtcc tggtccactt gacgaggaag ggaaccggaa accactatca gtatccacag
     5401 gaaatacagt tctatactcc aaatatgctg gcaatgaatt caaaggtgct gatggttctg
     5461 agtacattgc actgagggca tctgatgtaa tggctgtgct ttcatagggc gcgcgaagtt
     5521 aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag aaataatttt
     5581 gtttaacttt aagaaggaga tatacatacc catgtcacca caaacagaga ctaaagcaag
     5641 tgttggattc aaagctggtg ttaaagagta caaattgact tattatactc ctgagtacca
     5701 aaccaaggat actgatatat tggcagcatt ccgagtaact cctcaacctg gagttccacc
     5761 tgaagaagca ggggccgcgg tagctgccga atcttctact ggtacatgga caactgtatg
     5821 gaccgatgga cttaccagcc ttgatcgtta caaagggcga tgctaccgca tcgagcgtgt
     5881 tgttggagaa aaagatcaat atattgctta tgtagcttac cctttagacc tttttgaaga
     5941 aggttctgtt accaacatgt ttacttccat tgtaggtaac gtatttgggt tcaaagccct
     6001 gcgcgctcta cgtctggaag atctgcgaat ccctcctgct tatgttaaaa ctttccaagg
     6061 tccgcctcat gggatccaag ttgaaagaga taaattgaac aagtatggtc gtcccctgtt
     6121 gggatgtact attaaaccta aattggggtt atctgctaaa aactacggta gagctgttta
     6181 tgaatgtctt cgcggtggac ttgattttac caaagatgat gagaacgtga actcacaacc
     6241 atttatgcgt tggagagatc gtttcttatt ttgtgccgaa gcactttata aagcacaggc
     6301 tgaaacaggt gaaatcaaag ggcattactt gaatgctact gcaggtacat gcgaagaaat
     6361 gatcaaaaga gctgtatttg ctagagaatt gggcgttccg atcgtaatgc atgactactt
     6421 aacgggggga ttcaccgcaa atactagctt ggctcattat tgccgagata atggtctact
     6481 tcttcacatc caccgtgcaa tgcatgcggt tattgataga cagaagaatc atggtatcca
     6541 cttccgggta ttagcaaaag cgttacgtat gtctggtgga gatcatattc actctggtac
     6601 cgtagtaggt aaacttgaag gtgaaagaga cataactttg ggctttgttg atttactgcg
     6661 tgatgatttt gttgaacaag atcgaagtcg cggtatttat ttcactcaag attgggtctc
     6721 tttaccaggt gttctacccg tggcttcagg aggtattcac gtttggcata tgcctgctct
     6781 gaccgagatc tttggggatg attccgtact acagttcggt ggaggaactt taggacatcc
     6841 ttggggtaat gcgccaggtg ccgtagctaa tcgagtagct ctagaagcat gtgtaaaagc
     6901 tcgtaatgaa ggacgtgatc ttgctcagga aggtaatgaa attattcgcg aggcttgcaa
     6961 atggagcccg gaactagctg ctgcttgtga agtatggaaa gagatcgtat ttaattttgc
     7021 agcagtggac gttttggata agtaatactt ggcgcgccca gtgcggccgc ataatgctta
     7081 agtcgaacag aaagtaatcg tattgtacac ggccgcataa tcgaaattaa tacgactcac
     7141 tataggggaa ttgtgagcgg ataacaattc cccatcttag tatattagtt aagtataaga
     7201 aggagatata catatacata tggcagcaaa ggagttgtat ttcaacaagg atggatcagc
     7261 tatcaagagg ctgcaggttg gtgtgaacaa actcgctgat ctagttggag ttactcttgg
     7321 cccaaaaggc aggaatgttg ttcttgagag caagtatggt gctccaaaga ttgtcaatga
     7381 tggagttact gttgccagag aggttgagtt agaggatcca gtagaaaaca ttggtgctaa
     7441 actagtgaga caagctgctg ccaaaaccaa tgatttggct ggtgatggga ctacaacatc
     7501 tgtcgtacta gcccagggcc tggttgctga aggtgttaag gtggttgcag ctggtgcaaa
     7561 ccctgtcctg atcaccagag gaattgagaa gaccacaaaa gccttagtag ctgaattgaa
     7621 gaatatgtca aaagaggtag aagacagtga attagcagat gttgctgcag taagtgctgg
     7681 gaacaactct gaagtaggga gtatgattgc tgaagccatg agcaaggttg gaaggaaagg
     7741 tgttgtgact cttgaagagg gaaaaagtgc tgaaaacagt ctccgtgtgg ttgaaggaat
     7801 gcaatttgac cgtggttaca tctcaccata cttcgttact gatagtgaga aaatgtccgt
     7861 tgagtatgag aactgtaagt tattactggt tgacaagaag ataacaaatg caagagatct
     7921 tgttggtgtc ctggaagatg ctatcagaaa tggttaccca attttaatta ttgccgaaga
     7981 tattgagcaa gaagcactag ccacccttgt tgtcaataag cttagaggtg ctttaaaggt
     8041 tgctgccctt aaagctcctg gttttggtga acgaaaaagc cagtatcttg atgacatagc
     8101 aacccttact ggaggcactg tgattaggga ggaacttggc cttaccttag acaaggctga
     8161 caaggaagtt ctaggccatg cttctaaggt agtgctgact aaggatgcca ccacaattgt
     8221 tggtgatggt agcactcagg aagcagtcaa taagcgtgtt gtacagatta aaaacctcat
     8281 agaggcggca gatcaagatt atgaaaaaga aaagctaaat gaaagaattg ctaaattatc
     8341 aggaggtgtg gctgttatac aggttggagc acaaactgaa actgaattga aggaaaagaa
     8401 gctgagggtg gaagatgctc tcaatgcaac aaaggcagct gttgaggaag gtattgttgt
     8461 tggtggaggg tgcacactcc tgcgacttgc ttctaaagtt gatgccatca aggagactct
     8521 tgagaatgat gaggagaagg ttggagctga tattgttaaa agagcgttga gctacccatt
     8581 gaagttgatt gctaaaaatg ctggtgttaa tggaagtgtt gtcagtgaga aggtgctctc
     8641 tagtgacaat ccaaaatttg gatacaatgc tgcaactggc aattatgaag acttgatggc
     8701 tgctggtatt attgacccaa caaaggtggt gagatgttgt ctagagcatg cgtcatcagt
     8761 ggcgaagaca ttcttgatgt cagattgtgt ggtggtagag atcaaggagc ctgaaccagc
     8821 agttgctggc aacccaatgg acaattcagg ttatggatac taattatgtt atatctttgt
     8881 aatctgcaac gtaccgaaag gaagctgagt tggctgctgc caccgctgag caataactag
     8941 cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta
     9001 tatccggata tcccgcaaga ggcccggcag taccggcata accaagccta tgcctacagc
     9061 atccagggtg acggtgccga ggatgacgat gagcgcattg ttagatttca tacacggtgc
     9121 ctgactgcgt tagcaattta actgtgataa actaccgcat taaagctagc ttatcgatga
     9181 taagctgtca aacatgagaa t
//
