LOCUS       sequence-324149-        5986 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  p1A negative control expressing CbbLS and inactive Prk (Prk K20M 
            S21A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5986)
  AUTHORS   Flamholz AI, Dugan E, Blikstad C, Gleizer S, Ben-Nissan R, Amram S, 
            Antonovsky N, Ravishankar S, Noor E, Bar-Even A, Milo R, Savage D
  TITLE     Functional reconstitution of a bacterial CO2 concentrating mechanism
            in E. coli.
  JOURNAL   Elife. 2020 Oct 21;9. pii: 59882. doi: 10.7554/eLife.59882.
  PUBMED    33084575
REFERENCE   2  (bases 1 to 5986)
  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: "Elife."; 
            date: "2020-10-21"; volume: "9. pii: 59882. doi"; pages: " 
            10.7554/eLife.59882"
COMMENT     Sequence Label: p1A Prk-
FEATURES             Location/Qualifiers
     source          1..5986
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     267..286
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     519..537
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(558..576,577..582,583..599,600..605,606..631)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    558..576
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    583..601
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             641..652
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2501..2518
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3568..3654
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3827..4450)
                     /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     4358..4377
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(4451..4555)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4731..5319)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4811..4830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(5407..5501)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(5532..5986,1..340))
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5643..5662)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc acttcgccca
       61 atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagctgcg caaggaacgc
      121 ccgtcgtggc cagccacgat agccgcgctg cctcgtcctg cagttcattc agggcaccgg
      181 acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc tgacagccgg aacacggcgg
      241 catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc tccacccaag
      301 cggccggaga acctgcgtgc aatccatctt gttcaatcat gcgaaacgat cctcatcctg
      361 tctcttgatc agatcttgat cccctgcgcc atcagatcct tggcggcaag aaagccatcc
      421 agtttacttt gcagggcttc ccaaccttac cagagggcgc cccagctggc aattccgacg
      481 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
      541 ttcgtcttca cctcgagtcc ctatcagtga tagagattga catccctatc agtgatagag
      601 atactgagca catcagcagg acgcactgac cgaattcatt aaagaggaga aaggtaccat
      661 ggcagttaaa aagtatagtg ctggtgtaaa agaataccgg cagacctatt ggatgccgga
      721 atacacaccg ttggattccg acatccttgc atgcttcaaa atcaccccac aaccgggtgt
      781 tgatcgcgaa gaagccgcag ccgcggttgc agcagaatct tcaaccggca catggaccac
      841 cgtgtggacc gatttgctga ccgacatgga ctactacaaa ggccgtgcct atcgcattga
      901 agacgtaccc ggtgacgatg cggcattcta tgcctttatc gcctacccaa tcgacctgtt
      961 cgaagaaggg tcagttgtta acgtgtttac ctcactggtt ggtaacgtgt tcggcttcaa
     1021 agcggtacgc ggcctgcgtc tggaagatgt tcgcttccca ctcgcctacg ttaaaacctg
     1081 tggcggccca ccgcacggta ttcaagtcga acgcgacaag atgaacaaat atggtcgccc
     1141 actgttgggt tgcaccatca agccaaaact tggtttgtct gcgaaaaact acggccgtgc
     1201 cgtatacgag tgcctccgtg gcggcctcga cttcactaaa gatgatgaaa acatcaattc
     1261 tcagccgttc atgcgctggc gcgatcgctt cttgttcgta caagacgcga ccgaaactgc
     1321 tgaagcccaa accggcgaac gcaaaggcca ttacctcaac gtaacggcgc caactcctga
     1381 agaaatgtac aagcgcgcag aattcgccaa agaaattggc gcgccaatca ttatgcacga
     1441 ctacatcacc ggtggcttca cggccaacac tggcttggcc aagtggtgtc aagacaacgg
     1501 cgtactgctg cacatccacc gtgcgatgca tgcggttatc gaccgtaacc cgaaccacgg
     1561 tattcacttc cgtgttctga ccaagattct gcgtttgtcg ggtggcgatc acctgcacac
     1621 cggtaccgtt gtcggcaaac tggaaggcga ccgtgcctct actctgggct ggattgattt
     1681 gctccgcgaa tcgtttatcc ctgaagatcg ctcgcgcggt atcttcttcg atcaagactg
     1741 gggttcaatg ccaggcgtat tcgctgtggc ctctggtggt attcacgtat ggcacatgcc
     1801 tgcgctggtt aacatctttg gtgacgactc tgtcctccaa ttcggtggcg gtacgctggg
     1861 tcatccatgg ggcaacgctg ccggtgctgc tgccaaccgt gttgctctgg aagcctgcgt
     1921 agaagcgcgt aaccaaggcc gcgatatcga aaaagaaggc aaagaaattc tgactgctgc
     1981 tgcacagcac agcccagaac tgaagattgc catggaaact tggaaagaga tcaaattcga
     2041 atttgacact gtcgacaaac tcgacactca aaatcgttga tccctcgtac cacacaacat
     2101 actaaggtga gtaaccatgg ctgaaatgca ggattacaag caaagcctca aatatgagac
     2161 tttctcttat cttccaccca tgaacgcgga acgcatccgc gctcaaatca agtacgcaat
     2221 tgctcaaggc tggagccccg gcattgagca cgtagaagtg aaaaactcca tgaaccaata
     2281 ttggtacatg tggaaacttc ccttcttcgg cgaacaaaat gtcgacaacg tgttggctga
     2341 aattgaagcg tgtcgtagtg cgtatccaac acaccaggtc aaactggtgg cttatgacaa
     2401 ctatgcgcaa agcttaggtc tggccttcgt ggtctaccgc ggcaactaaa tctaatagaa
     2461 ataattttgt ttaactttat aagcaggacc ggcggcgatg catcatcacc atcaccacag
     2521 caagccagat cgtgttgttt tgatcggcgt tgccggtgac tccggttgcg gcatggcaac
     2581 cttcctaaat cgccttgccg acttgtttgg tacggaattg atgacggtca tctgcttgga
     2641 tgactatcac agtctcgatc gcaagggccg gaaggaagca ggcgtaacgg ctttggatcc
     2701 ccgcgccaac aactttgact tgatgtatga acaggtcaag gcgttgaaga acggcgaaac
     2761 gatcatgaag ccgatctaca accatgaaac cggcttgatc gatccgcccg aaaaaatcga
     2821 acccaatcgc atcattgtga tcgagggtct gcatccgctt tacgacgagc gcgtgcgtga
     2881 actgctcgat ttcagcgttt acctcgacat cgatgacgaa gtcaaaatcg cttggaagat
     2941 ccaacgcgat atggcagaac gcggccactc ctacgaagat gtcctcgcct cgatcgaagc
     3001 gcgccgccct gacttcaagg cctacattga gccccagcgt ggccatgcgg acatcgtcat
     3061 ccgcgtcatg ccgacccagc taatccccaa tgacaccgag cgcaaggtgc tgcgggtgca
     3121 gttgatccaa cgggaaggcc gcgatggttt tgagccggct tacctgttcg acgaaggttc
     3181 gaccatccag tggacgccct gcggtcgtaa gctgacctgc tcctatccgg gcattcgctt
     3241 agcctacggc cctgacacct actacggtca cgaagtctca gtgcttgagg tcgacggtca
     3301 gttcgagaac ctcgaagaga tgatctacgt cgagggccac ctcagcaaga ccgacacgca
     3361 gtactacggt gagttgaccc acctgctgct acagcacaaa gattacccgg gttcgaacaa
     3421 cggcacgggt ctgttccaag tgctgaccgg cctgaaaatg cgggcggcct atgagcgttt
     3481 gacctcccaa gcagcacccg tcgccgctag tgtctagatg gaattcctgc agcccggggg
     3541 atcccatggt acgcgtgcta gaggcatcaa ataaaacgaa aggctcagtc gaaagactgg
     3601 gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac aaatccgccg
     3661 ccgctagcag ttcgtagacg gaaacaaacg cagaatccaa gcaaaaggga ataagggcga
     3721 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     3781 gttatgtcca ctgggttcgt gccttcatcc ggatatccag gaactgttaa gacccacttt
     3841 cacatttaag ttgtttttct aatccgcata tgatcaattc aaggccgaat aagaaggctg
     3901 gctctgcacc ttggtgatca aataattcga tagcttgtcg taataatggc ggcatactat
     3961 cagtagtagg tgtttccctt tcttctttag cgacttgatg ctcttgatct tccaatacgc
     4021 aacctaaagt aaaatgcccc acagcgctga gtgcatataa tgcattctct agtgaaaaac
     4081 cttgttggca taaaaaggct aattgatttt cgagagtttc atactgtttt tctgtaggcc
     4141 gtgtacctaa atgtactttt gctccatcgc gatgacttag taaagcacat ctaaaacttt
     4201 tagcgttatt acgtaaaaaa tcttgccagc tttccccttc taaagggcaa aagtgagtat
     4261 ggtgcctatc taacatctca atggctaagg cgtcgagcaa agcccgctta ttttttacat
     4321 gccaatacaa tgtaggctgc tctacaccta gcttctgggc gagtttacgg gttgttaaac
     4381 cttcgattcc gacctcatta agcagctcta atgcgctgtt aatcacttta cttttatcta
     4441 atctagacat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4501 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgtttcc
     4561 ctcgacaatt cgcgctagac ctagggcgtt cggctgcggc gagcggtatc agctcactca
     4621 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4681 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4741 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4801 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4861 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4921 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4981 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5041 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5101 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5161 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5221 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5281 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5341 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgactagt
     5401 gcttggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga acaaatccag
     5461 atggagttct gaggtcatta ctggatctat caacaggagt ccaagcgagc tctcgaaccc
     5521 cagagtcccg ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg
     5581 ggagcggcga taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca
     5641 gcaatatcac gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca
     5701 cagtcgatga atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg
     5761 ccatgggtca cgacgagatc ctcgccgtcg ggcatgcgcg ccttgagcct ggcgaacagt
     5821 tcggctggcg cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct
     5881 tccatccgag tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta
     5941 gccggatcaa gcgtatgcag ccgccgcatt gcatcagcca tgatgg
//
