LOCUS       sequence-467655-        6272 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing rubisco and prk in Delta-rpi.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6272)
  AUTHORS   Prywes N, Phillips NR, Oltrogge LM, Lindner S, Taylor-Kearney LJ, 
            Tsai YC, de Pins B, Cowan AE, Chang HA, Wang RZ, Hall LN, 
            Bellieny-Rabelo D, Nisonoff HM, Weissman RF, Flamholz AI, Ding D, 
            Bhatt AY, Mueller-Cajar O, Shih PM, Milo R, Savage DF
  TITLE     A map of the rubisco biochemical landscape.
  JOURNAL   Nature. 2025 Jan 22. doi: 10.1038/s41586-024-08455-0.
  PUBMED    39843747
REFERENCE   2  (bases 1 to 6272)
  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: "Nature. 
            2025 Jan 22. doi: 10.1038/s41586-024-08455-0."
COMMENT     Sequence Label: NP-11-64-1
FEATURES             Location/Qualifiers
     source          1..6272
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1832..1850
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1857..1875
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        1888..1992
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1993..2616
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(2066..2085)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        2623..2725
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2726..3385
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2792..2811)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3397..3491
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      3605..4150
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4051..4070
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4267..4284
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(4387..5469)
                     /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     5431..5450
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5470..5547)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(5591..5596,5597..5612,5613..5619)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    5625..5644
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     RBS             5677..5685
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
ORIGIN
        1 gcaagggaac caacccttcg cgccacttcg cgacacgatc gccttggttg cagatgcgat
       61 gcgccgtgca caggacgaaa cgggtgaagc aaagttgttt agtgctaata tcaccgctga
      121 tgacccgttt gaaatcatcg cccgcgggga atacgttctt gaaacattcg gtgaaaatgc
      181 aagtcacgtt gccttattag tagacggata cgtagccggc gcagctgcta ttacgaccgc
      241 tcgtcgccgt ttccccgata atttcttgca ttatcatcgc gccgggcatg gagcggtcac
      301 tagccctcag tctaagcgcg gctatactgc ttttgtgcat tgtaagatgg ctcgtttgca
      361 aggtgcgtcg ggtatccata ccggaactat gggctttgga aagatggaag gggagtcctc
      421 agatcgtgcc atcgcgtata tgttaactca agatgaagca caaggcccct tttatcgcca
      481 gtcgtgggga gggatgaagg cctgcacccc tattatttct ggcggaatga atgccttacg
      541 catgcccggc ttcttcgaaa atcttggtaa tgcgaatgtc atcctgactg ctggtggcgg
      601 tgctttcggc catattgacg gccctgtggc cggcgctcgt tctcttcgcc aagcttggca
      661 ggcgtggcgc gatggcgtcc ctgtgcttga ctacgcccgc gagcataagg agttggcacg
      721 tgcttttgag tccttccctg gcgacgctga ccagatttat ccaggttggc gtaaggcgct
      781 tggagtggag gatactcgtt cagctttacc tgcgtaatag cctgcaggac cggtggtgac
      841 ctcgcagcat agattaccat gcttaagcaa ttgaacgaga aaagccaacc tgcgggttgg
      901 cttttttatg caacgcgttt actgcgcggc gcgcttgcgg cgctccatca tgcgcagcac
      961 gaagggcgtg aagatcagct gcatggccag ctccatcttg ccgccgggca ccacgatggt
     1021 gttggcgcgc gacatgaagg agtcgtggat catgctcagc aggtactgga agtcgatccc
     1081 cttcgggttg gcaaagcgga tcaccaccat gctttcatcg ggcgcgggga tttcgcgcga
     1141 gataaagggg ttggacgtat ccacgcacgg cacgcgctgg aagttcacat gcgtgcgcga
     1201 gaactgcgga cagatgtaat tcacgtagtc cggcatgcgg cgcaggatgg tgtcggtcac
     1261 ggcctcggtc gaatagccgc gctgcttctt gtcgcgccag agcttctgga tccattccag
     1321 gttgatgaca gggaccacgc cgatcagcag gttgggatac tgggcgacat tgaccgaatc
     1381 ggtgaccacg ccgccgtgca ggccctcgta gaacagcaga tcggtatcgg ccggcagcgg
     1441 ctcccactgc gtgaaggtgc cgggttcctg cccgaacggc gcggcttcct ccgggctatg
     1501 caggtagtgc cggtgcatgc ccgtgccggt ttcggcatag gagcggaaca ggttctccag
     1561 ctcgccgaac aggttgttct cctcgccgaa gtggctgaag ttcatattgc cggtgcgctc
     1621 ggcctcggcc atcttgacct tcatctcggc gcggtcatag cggtggaagc tgtcgccttc
     1681 gatcaccact gacttcacgc cctcgcggcg gaagatgttc tcgaaggtac gggtcaccga
     1741 cgtggttccg gcgcccgagg agccggtgat ggcgataatg ggataacgtt ctgacataaa
     1801 aaacctccta gcacattatc agtgctcagt atctctatca ctgataggga tgtcaatctc
     1861 tatcactgat agggactcga ggggccccgc ggaaccccta tttgtttatt tttctaaata
     1921 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     1981 aaaaggaaga gtatgtctag attagataaa agtaaagtga ttaacagcgc attagagctg
     2041 cttaatgagg tcggaatcga aggtttaaca acccgtaaac tcgcccagaa gctaggtgta
     2101 gagcagccta cattgtattg gcatgtaaaa aataagcggg ctttgctcga cgccttagcc
     2161 attgagatgt tagataggca ccatactcac ttttgccctt tagaagggga aagctggcaa
     2221 gattttttac gtaataacgc taaaagtttt agatgtgctt tactaagtca tcgcgatgga
     2281 gcaaaagtac atttaggtac acggcctaca gaaaaacagt atgaaactct cgaaaatcaa
     2341 ttagcctttt tatgccaaca aggtttttca ctagagaatg cattatatgc actcagcgct
     2401 gtggggcatt ttactttagg ttgcgtattg gaagatcaag agcatcaagt cgctaaagaa
     2461 gaaagggaaa cacctactac tgatagtatg ccgccattat tacgacaagc tatcgaatta
     2521 tttgatcacc aaggtgcaga gccagccttc ttattcggcc ttgaattgat catatgcgga
     2581 ttagaaaaac aacttaaatg tgaaagtggg tcttaagacg tctgatcggc acgtaagagg
     2641 ttccaacttt caccataatg aaataagatc actaccgggc gtattttttg agttatcgag
     2701 attttcagga gctaaggaag ctaaaatgga gaaaaaaatc actggatata ccaccgttga
     2761 tatatcccaa tggcatcgta aagaacattt tgaggcattt cagtcagttg ctcaatgtac
     2821 ctataaccag accgttcagc tggatattac ggccttttta aagaccgtaa agaaaaataa
     2881 gcacaagttt tatccggcct ttattcacat tcttgcccgc ctgatgaatg ctcatccgga
     2941 attccgtatg gcaatgaaag acggtgagct ggtgatatgg gatagtgttc acccttgtta
     3001 caccgttttc catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga
     3061 tttccggcag tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc
     3121 ctatttccct aaagggttta ttgagaatat gtttttcgtc tcagccaatc cctgggtgag
     3181 tttcaccagt tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac
     3241 catgggcaaa tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca
     3301 tcatgccgtt tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg
     3361 cgatgagtgg cagggcgggg cgtaagagct cgcttggact cctgttgata gatccagtaa
     3421 tgacctcaga actccatctg gatttgttca gaacgctcgg ttgccgccgg gcgtttttta
     3481 ttggtgagaa tccaagcact agtaacaact tatatcgtat ggggctgact tcaggtgcta
     3541 catttgaaga gataaattgc actgaaatct agaaatattt tatctgatta ataagatgat
     3601 cttcttgaga tcgttttggt ctgcgcgtaa tctcttgctc tgaaaacgaa aaaaccgcct
     3661 tgcagggcgg tttttcgaag gttctctgag ctaccaactc tttgaaccga ggtaactggc
     3721 ttggaggagc gcagtcacca aaacttgtcc tttcagttta gccttaaccg gcgcatgact
     3781 tcaagactaa ctcctctaaa tcaattacca gtggctgctg ccagtggtgc ttttgcatgt
     3841 ctttccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc ggactgaacg
     3901 gggggttcgt gcatacagtc cagcttggag cgaactgcct acccggaact gagtgtcagg
     3961 cgtggaatga gacaaacgcg gccataacag cggaatgaca ccggtaaacc gaaaggcagg
     4021 aacaggagag cgcacgaggg agccgccagg gggaaacgcc tggtatcttt atagtcctgt
     4081 cgggtttcgc caccactgat ttgagcgtca gatttcgtga tgcttgtcag gggggcggag
     4141 cctatggaaa aacggctttg ccgcggccct ctcacttccc tgttaagtat cttcctggca
     4201 tcttccagga aatctccgcc ccgttcgtaa gccatttccg ctcgccgcag tcgaacgacc
     4261 gagcgtagcg agtcagtgag cgaggaagcg gaatatatcc ctaggagaaa tcatccttag
     4321 cgaaagctaa ggattttttt tatctggctg ggagttcgta gacggaaaca aacgcagaat
     4381 ccaagctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4441 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     4501 cagtgaaacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     4561 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     4621 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     4681 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     4741 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     4801 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     4861 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     4921 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     4981 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     5041 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     5101 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     5161 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     5221 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     5281 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     5341 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     5401 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     5461 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     5521 aaaggttttg cgccattcga tggtgtcgca ctgaaggtcc tcaatcgcac tggaaacatc
     5581 aaggtcggtg ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcgctcac
     5641 aattatttca ctggaaatta cgccagacat ccctaataag gaggtaccat ggaccagagt
     5701 tctcgctatg tcaatcttgc tttgaaagaa gaggacttaa ttgccggagg cgaacacgta
     5761 ttgtgcgcct atattatgaa accaaaagct gggtacggtt acgttgccac cgcagcgcac
     5821 tttgctgcag agagttcgac tggaacaaat gtagaggtgt gtaccactga tgattttact
     5881 cgtggcgtgg atgccttagt ttacgaagta gatgaggccc gcgagcttac taagattgcc
     5941 tacccagtag cactgttcga tcgtaatatc acggatggaa aagctatgat cgcatcattt
     6001 cttacattga ccatgggcaa taatcaaggc atgggtgacg tagaatatgc caaaatgcat
     6061 gacttctatg tgcctgaagc gtatcgcgct ctgtttgatg gcccttctgt aaatatttcc
     6121 gccttgtgga aggtactggg tcgtccggag gttgatggag gcttagttgt cggtactatc
     6181 attaaaccaa agcttgggct gcgccctaag cctttcgctg aggcttgcca cgcattttgg
     6241 ttaggtggtg attttattaa aaatgatgaa cc
//
