LOCUS       sequence-472475-        7506 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses G. max RbcL RbcS-6xHis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7506)
  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 7506)
  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: pTW335f3
FEATURES             Location/Qualifiers
     source          1..7506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     170..189
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     423..440
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    455..594
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(520..539)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     680..702
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(696..887)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2199..3281)
                     /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     3243..3262
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3282..3359)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(3602..3607,3608..3623,3624..3630)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    3638..3654
                     /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     3643..3665
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     RBS             5172..5194
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5586..5603
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      5684..5731
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(5965..5983)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6051..6155
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6156..6224,6225..7016)
                     /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(6374..6393)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7187..7506,1..269)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
       61 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
      121 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
      181 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
      241 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
      301 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
      361 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
      421 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
      481 atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc
      541 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
      601 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
      661 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
      721 aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg
      781 tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt
      841 ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg
      901 taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg
      961 atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg
     1021 gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt
     1081 aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac
     1141 ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag
     1201 accattcatg ttgttgctca ggtcgcagat gttttgcagc agcagtcgct tcacgttcgc
     1261 tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc
     1321 ctcaacgaca ggagcacgat catgcgcacc cgtggccagg acccaacgct gcccgagatg
     1381 cgccgcgtgc ggctgctgga gatggcggac gcgatggata tgttctgcca agggttggtt
     1441 tgcgcattca cagttctccg caagaattga ttggctccaa ttcttggagt ggtgaatccg
     1501 ttagcgaggt gccgccggct tccattcagg tcgaggtggc ccggctccat gcaccgcgac
     1561 gcaacgcggg gaggcagaca aggtataggg cggcgcctac aatccatgcc aacccgttcc
     1621 atgtgctcgc cgaggcggca taaatcgccg tgacgatcag cggtccaatg atcgaagtta
     1681 ggctggtaag agccgcgagc gatccttgaa gctgtccctg atggtcgtca tctacctgcc
     1741 tggacagcat ggcctgcaac gcgggcatcc cgatgccgcc ggaagcgaga agaatcataa
     1801 tggggaaggc catccagcct cgcgtcgcga acgccagcaa gacgtagccc agcgcgtcgg
     1861 ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg ggaccagtga
     1921 cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg ccgatcatcg
     1981 tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc
     2041 ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc atgccccgcg
     2101 cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg
     2161 cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg
     2221 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     2281 gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc
     2341 ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg
     2401 cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg
     2461 tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt aatggcgcgc
     2521 attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca
     2581 ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc
     2641 gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc
     2701 gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc
     2761 agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt
     2821 gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca
     2881 atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga
     2941 agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc
     3001 acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg
     3061 tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt
     3121 tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc
     3181 gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca
     3241 ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga
     3301 ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcg
     3361 cgcaacgcaa ttaatgtaag ttagctcact cattaggcac aattctcatg tttgacagct
     3421 tatcatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg
     3481 tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt
     3541 ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct
     3601 gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata acaatttcac
     3661 acaggaaaca gccagtccgt ttaggtgttt tcacgagcac ttcaccaaca ataggtgcag
     3721 taaggagata taccatgcca caaacagaga ctaaagcaag tgttgggttc aaagctggtg
     3781 ttaaagatta taaattgact tattatactc ctgactatga aaccaaagat actgatatct
     3841 tggcagcatt ccgagtaact cctcaaccag gagttccgcc tgaagaagca ggtgccgcgg
     3901 tagccgccga atcttctact ggtacatgga caactgtgtg gaccgatggg cttaccagtc
     3961 ttgatcgtta caaagggcga tgctacggcc ttgaacctgt tgctggggaa gaaaatcaat
     4021 atattgctta tgtagcttat cccttagacc tttttgaaga aggttctgtt actaacatgt
     4081 ttacttccat tgtcggtaat gtatttgggt tcaaggccct gcgtgctcta cgtctggagg
     4141 atttgcgaat ccctactgct tatattaaaa ctttccaagg tccgcctcat ggcatccaag
     4201 ttgagagaga taaattgaac aagtatggtc gtcccctatt aggatgtact attaaaccta
     4261 aattggggtt atccgctaag aattatggta gagctgttta tgaatgtctt cgtgggggac
     4321 ttgattttac caaagatgat gaaaatgtga attcccaacc atttatgcgt tggagagacc
     4381 gtttcttatt ttgtgccgaa gccattttta aatcacaggc tgaaacaggt gaaatcaaag
     4441 ggcattactt gaatgcaact gcgggtacat gcgaagaaat gatgaaaaga gctgtatttg
     4501 ccagagaatt aggcgttcct atcgtaatgc atgattattt aacgggggga ttcactgcaa
     4561 atactagctt ggctcattat tgccgagata atggtctact tcttcatata caccgtgcaa
     4621 tgcatgcagt tatcgacaga caaaagaatc atggtatgca ctttcgtgta ctagctaaag
     4681 cattacgttt gtctggtgga gatcatgttc acgccggtac cgtagtaggt aaacttgaag
     4741 gggaaagaga aatcacttta ggttttgttg atttactacg tgatgatttt gttgaaaaag
     4801 atcgaagtcg cggtatttat ttcactcagg attgggtttc tctaccaggt gttttgcctg
     4861 tagcttcggg aggtattcac gtttggcata tgcctgctct gaccgagatc tttggggatg
     4921 attctgtact ccaatttggc ggaggaactt taggacaccc ttggggaaat gcaccaggtg
     4981 ctgtagctaa tcgagtagct cttgaagcat gtgtgcaggc tcgaaatgaa ggacgcgatc
     5041 ttgctcgtga aggtaatgaa attatccgtg aggctagcaa atggagtcct gaattagctg
     5101 ctgcttgtga agtatggaag gagatcaaat ttgaattcga agcaatggat actttgtaag
     5161 ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgtgtatg caagtttggc
     5221 cccccatcgg taagaaaaaa tttgagaccc tgtcatactt gccagacctg gatgatgccc
     5281 agttagctaa agaagtggaa tatcttctcc gaaaaggttg gattccgtgt ctggaatttg
     5341 agttagagca tggcttcgtt taccgcgaac acaaccgtag tccggggtat tatgacggcc
     5401 gttactggac tatgtggaaa ctgcctatgt tcggatgcac cgatgcgagc caggtcctaa
     5461 aagaactgca agaagcgaag acggcctatc cgaacggctt catccgcatt attggctttg
     5521 ataatgtgcg ccaggtacaa tgtatttcgt ttatagcata caaaccgcct ggttttgcac
     5581 tcgagcatca ccaccatcat cattaaatcc ggctgctaac aaagcccgaa aggaagctga
     5641 gttggctgct gccaccgctg agcaataaac cgctgagcaa taactagcat aaccccttgg
     5701 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggatatcc
     5761 cgcaagaggc ccggcagtac cggcataacc aagcctatgc ctacagcatc cagggtgacg
     5821 gtgccgagga tgacgatgag cgcattgtta gatttcatac acggtgcctg actgcgttag
     5881 caatttaact gtgataaact acagcattaa agctagctta tcgatgataa gctgtcaaac
     5941 atgagaattc ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     6001 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     6061 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     6121 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     6181 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     6241 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     6301 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     6361 cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac
     6421 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     6481 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6541 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6601 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6661 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
     6721 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6781 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6841 ttgctgataa atctggagcc ggtgagcgtg gatctcgcgg tatcattgca gcactggggc
     6901 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6961 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     7021 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     7081 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     7141 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     7201 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     7261 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     7321 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     7381 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     7441 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     7501 gctgaa
//
