LOCUS       sequence-472477-        7522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses A. thaliana RbcL RbcS-6xHis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7522)
  AUTHORS   Archer J, Kathpalia M, Lee BU, Li S, Wang T
  TITLE     Effects of chaperone selectivity on the assembly of plant Rubisco 
            orthologs in E. coli.
  JOURNAL   J Exp Bot. 2025 Mar 31:eraf140. doi: 10.1093/jxb/eraf140.
  PUBMED    40163664
REFERENCE   2  (bases 1 to 7522)
  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: "J Exp Bot. 
            2025 Mar 31:eraf140. doi: 10.1093/jxb/eraf140."
COMMENT     Sequence Label: pTW335c3
FEATURES             Location/Qualifiers
     source          1..7522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1..1083)
                     /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     1045..1064
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1084..1161)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(1404..1409,1410..1425,1426..1432)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1440..1456
                     /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     1445..1467
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     RBS             2986..3008
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3403..3420
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3502..3549
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(3783..3801)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3869..3973
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3974..4042,4043..4834)
                     /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(4192..4211)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5005..5593
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5494..5513
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5747..5764
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5779..5918
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5844..5863)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6004..6026
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6020..6211)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
       61 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
      121 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
      181 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
      241 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag
      301 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
      361 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
      421 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
      481 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
      541 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
      601 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
      661 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
      721 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
      781 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
      841 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
      901 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
      961 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     1021 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     1081 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     1141 tttgcgccat tcgatggtgt cgcgcaacgc aattaatgta agttagctca ctcattaggc
     1201 acaattctca tgtttgacag cttatcatcg actgcacggt gcaccaatgc ttctggcgtc
     1261 aggcagccat cggaagctgt ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg
     1321 tcgctcaagg cgcactcccg ttctggataa tgttttttgc gccgacatca taacggttct
     1381 ggcaaatatt ctgaaatgag ctgttgacaa ttaatcatcg gctcgtataa tgtgtggaat
     1441 tgtgagcgga taacaatttc acacaggaaa cagccagtcc gtttaggtgt tttcacgagc
     1501 acttcaccaa caataggtgc agtaaggaga tataccatgc cacaaacaga gactaaagca
     1561 agtgttgggt tcaaagctgg tgttaaagag tataaattga cttactatac tcctgaatat
     1621 gaaaccaagg atactgatat cttggcagca ttccgagtaa ctcctcaacc tggagttcca
     1681 cctgaagaag caggggctgc ggtagctgct gaatcttcta ctggtacatg gacaactgtg
     1741 tggaccgatg ggcttaccag ccttgatcgt tacaaaggac gatgctacca catcgagccc
     1801 gttccaggag aagaaactca atttattgcg tatgtagctt atcccttaga cctttttgaa
     1861 gaaggttcgg ttactaacat gtttacctcg attgtgggta atgtatttgg gttcaaagcc
     1921 ctggctgctc tacgtctaga ggatctgcga atccctcctg cttatactaa aactttccaa
     1981 ggaccacctc atggtatcca agttgaaaga gataaattga acaagtatgg acgtccccta
     2041 ttaggatgta ctattaaacc aaaattgggg ttatccgcga aaaactatgg tagagcagtt
     2101 tatgaatgtc tacgtggtgg acttgatttt accaaagatg atgagaatgt gaactcccaa
     2161 ccatttatgc gttggagaga tcgtttctta ttttgtgccg aagctattta taaatcacag
     2221 gctgaaacag gtgaaatcaa agggcattat ttgaatgcta ctgcgggtac atgcgaagaa
     2281 atgatcaaaa gagctgtatt tgccagagaa ttgggagttc ctatcgtaat gcatgactac
     2341 ttaacagggg gattcaccgc aaatactagt ttgtctcatt attgccgaga taatggccta
     2401 cttcttcaca tccaccgtgc aatgcacgct gttattgata gacagaagaa tcatggtatg
     2461 cacttccgtg tactagctaa agctttacgt ctatctggtg gagatcatat tcacgcgggt
     2521 acagtagtag gtaaacttga aggagacagg gagtcaactt tgggctttgt tgatttactg
     2581 cgcgatgatt atgttgaaaa agatcgaagc cgcggtatct ttttcactca agattgggtt
     2641 tcactacctg gtgttctgcc tgtggcttca gggggtattc acgtttggca catgcctgct
     2701 ttgaccgaga tctttggaga tgattctgta ctacaattcg gtggaggaac tttaggccac
     2761 ccttggggaa atgcaccggg tgccgtagcc aaccgagtag ctctggaagc atgtgtacaa
     2821 gctcgtaatg agggacgtga tcttgcagtc gagggtaatg aaattatccg tgaagcttgc
     2881 aaatggagtc ctgaactagc tgctgcttgt gaagtatgga aagagatcac atttaacttc
     2941 ccaaccatcg ataaattaga tggccaagag taagctagaa ataattttgt ttaactttaa
     3001 gaaggagata tacatatgaa ggtgtggcca ccaattggaa agaagaagtt tgagactcta
     3061 tcttacctcc ctgaccttag tgacgtcgaa ttggctaagg aagttgacta ccttctccgc
     3121 aacaagtgga ttccttgtgt tgaattcgag ttagagcacg gatttgtgta ccgtgagcac
     3181 ggaaacactc ccggatacta cgatggacgg tactggacaa tgtggaagct tccattgttc
     3241 ggatgcaccg actccgctca agtgttgaag gaagttgaag aatgcaagaa ggagtacccg
     3301 ggcgccttca ttaggatcat cggattcgac aacacccgtc aagtccaatg catcagtttc
     3361 attgcctaca agcccccaag cttcaccgaa gctgcactcg agcaccacca ccaccaccac
     3421 tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     3481 caataaaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     3541 ggttttttgc tgaaaggagg aactatatcc ggatatcccg caagaggccc ggcagtaccg
     3601 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
     3661 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
     3721 cgcattaaag ctagcttatc gatgataagc tgtcaaacat gagaattctt gaagacgaaa
     3781 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     3841 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     3901 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     3961 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     4021 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     4081 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     4141 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     4201 cgcggtatta tcccgtgttg acgccgggca agagcaactc ggtcgccgca tacactattc
     4261 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     4321 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     4381 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     4441 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     4501 tgacaccacg atgcctgcag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     4561 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     4621 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     4681 tgagcgtgga tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     4741 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     4801 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     4861 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     4921 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     4981 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     5041 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     5101 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
     5161 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     5221 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     5281 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     5341 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     5401 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     5461 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     5521 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     5581 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     5641 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     5701 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     5761 cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     5821 acaccgcaat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     5881 acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg
     5941 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     6001 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc
     6061 tgcggtaaag ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg
     6121 cgtccagctc gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca
     6181 tgttaagggc ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt
     6241 tcatgggggt aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg
     6301 atgaacatgc ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc
     6361 gggaccagag aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg
     6421 ttccacaggg tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg
     6481 ctgacttccg cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg
     6541 ctcaggtcgc agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt
     6601 cattctgcta accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca
     6661 cgatcatgcg cacccgtggc caggacccaa cgctgcccga gatgcgccgc gtgcggctgc
     6721 tggagatggc ggacgcgatg gatatgttct gccaagggtt ggtttgcgca ttcacagttc
     6781 tccgcaagaa ttgattggct ccaattcttg gagtggtgaa tccgttagcg aggtgccgcc
     6841 ggcttccatt caggtcgagg tggcccggct ccatgcaccg cgacgcaacg cggggaggca
     6901 gacaaggtat agggcggcgc ctacaatcca tgccaacccg ttccatgtgc tcgccgaggc
     6961 ggcataaatc gccgtgacga tcagcggtcc aatgatcgaa gttaggctgg taagagccgc
     7021 gagcgatcct tgaagctgtc cctgatggtc gtcatctacc tgcctggaca gcatggcctg
     7081 caacgcgggc atcccgatgc cgccggaagc gagaagaatc ataatgggga aggccatcca
     7141 gcctcgcgtc gcgaacgcca gcaagacgta gcccagcgcg tcggccgcca tgccggcgat
     7201 aatggcctgc ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag
     7261 ggcgtgcaag attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa
     7321 gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat
     7381 aaagaagaca gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct
     7441 gactgggttg aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta
     7501 acttacatta attgcgttgc gc
//
