LOCUS       sequence-258480-        7659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CasA plant transcriptional activator (AT-CBF1) linked constitutive 
            expression cassette for Zea mays.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7659)
  AUTHORS   Young JK, Gasior SL, Jones S, Wang L, Navarro P, Vickroy B, 
            Barrangou R
  TITLE     The repurposing of type I-E CRISPR-Cascade for gene activation in 
            plants.
  JOURNAL   Commun Biol. 2019 Oct 18;2:383. doi: 10.1038/s42003-019-0637-6. 
            eCollection 2019.
  PUBMED    31646186
REFERENCE   2  (bases 1 to 7659)
  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: "Commun 
            Biol."; date: "2019-10-18"; volume: "2:383. doi"; pages: " 
            10.1038/s42003-019-0637-6. eCollection 2019"
COMMENT     Sequence Label: PV504
FEATURES             Location/Qualifiers
     source          1..7659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     7..26
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7..25
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        60..2055
                     /label=Ubi promoter
                     /note="maize polyubiquitin gene promoter"
     CDS             3751..3771
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        complement(4463..4481)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4464..4481)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4499..4515)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4499..4515)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4512..4534)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4523..4539
                     /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)."
     promoter        complement(join(4547..4553,4554..4571,4572..4577))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4592..4613
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4730..4747)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4901..5489)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4981..5000)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5663..5682)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5674..6489
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             5674..6489
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(5747..5766)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6783..6801
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6839..6861)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6961..6980
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(7047..7502)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7184..7205)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7396..7415
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7628..7650
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7642..7659
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7643..7659
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gaattgtaat acgactcact atagggcgaa ttgggttacc cggaccgaag cttgcatgcc
       61 tgcagtgcag cgtgacccgg tcgtgcccct ctctagagat aatgagcatt gcatgtctaa
      121 gttataaaaa attaccacat attttttttg tcacacttgt ttgaagtgca gtttatctat
      181 ctttatacat atatttaaac tttactctac gaataatata atctatagta ctacaataat
      241 atcagtgttt tagagaatca tataaatgaa cagttagaca tggtctaaag gacaattgag
      301 tattttgaca acaggactct acagttttat ctttttagtg tgcatgtgtt ctcctttttt
      361 tttgcaaata gcttcaccta tataatactt catccatttt attagtacat ccatttaggg
      421 tttagggtta atggttttta tagactaatt tttttagtac atctatttta ttctatttta
      481 gcctctaaat taagaaaact aaaactctat tttagttttt ttatttaata atttagatat
      541 aaaatagaat aaaataaagt gactaaaaat taaacaaata ccctttaaga aattaaaaaa
      601 actaaggaaa catttttctt gtttcgagta gataatgcca gcctgttaaa cgccgtcgac
      661 gagtctaacg gacaccaacc agcgaaccag cagcgtcgcg tcgggccaag cgaagcagac
      721 ggcacggcat ctctgtcgct gcctctggac ccctctcgag agttccgctc caccgttgga
      781 cttgctccgc tgtcggcatc cagaaattgc gtggcggagc ggcagacgtg agccggcacg
      841 gcaggcggcc tcctcctcct ctcacggcac cggcagctac gggggattcc tttcccaccg
      901 ctccttcgct ttcccttcct cgcccgccgt aataaataga caccccctcc acaccctctt
      961 tccccaacct cgtgttgttc ggagcgcaca cacacacaac cagatctccc ccaaatccac
     1021 ccgtcggcac ctccgcttca aggtacgccg ctcgtcctcc cccccccccn tctctacctt
     1081 ctctagatcg gcgttccggt ccatgcatgg ttagggcccg gtagttctac ttctgttcat
     1141 gtttgtgtta gatccgtgtt tgtgttagat ccgtgctgct agcgttcgta cacggatgcg
     1201 acctgtacgt cagacacgtt ctgattgcta acttgccagt gtttctcttt ggggaatcct
     1261 gggatggctc tagccgttcc gcagacggga tcgatttcat gatttttttt gtttcgttgc
     1321 atagggtttg gtttgccctt ttcctttatt tcaatatatg ccgtgcactt gtttgtcggg
     1381 tcatcttttc atgctttttt ttgtcttggt tgtgatgatg tggtctggtt gggcggtcgt
     1441 tctagatcgg agtagaattc tgtttcaaac tacctggtgg atttattaat tttggatctg
     1501 tatgtgtgtg ccatacatat tcatagttac gaattgaaga tgatggatgg aaatatcgat
     1561 ctaggatagg tatacatgtt gatgcgggtt ttactgatgc atatacagag atgctttttg
     1621 ttcgcttggt tgtgatgatg tggtgtggtt gggcggtcgt tcattcgttc tagatcggag
     1681 tagaatactg tttcaaacta cctggtgtat ttattaattt tggaactgta tgtgtgtgtc
     1741 atacatcttc atagttacga gtttaagatg gatggaaata tcgatctagg ataggtatac
     1801 atgttgatgt gggttttact gatgcatata catgatggca tatgcagcat ctattcatat
     1861 gctctaacct tgagtaccta tctattataa taaacaagta tgttttataa ttattttgat
     1921 cttgatatac ttggatgatg gcatatgcag cagctatatg tggatttttt tagccctgcc
     1981 ttcatacgct atttatttgc ttggtactgt ttcttttgtc gatgctcacc ctgttgtttg
     2041 gtgttacttc tgcaggtcga ctctagagga tccatgagcc gcttcaacct cctcgacgag
     2101 ccctggatca gcgtcatcta cgacgagaag ggcagcacca aggacgtcag cctccaggac
     2161 ctcttcacca acgcccacca gtacaaggag ctcgccggcg acaccaagac ccaggacttc
     2221 gccgtcctca gggtcctcct cgccgtcctc cacaccgtgt tcagccgctt cgacatcgac
     2281 ggcaacgcct acgagtacct caccatcgac gagggctgga accagctcga gcccgtcgac
     2341 gagatggaca tcgagaacta cgaggaggcc ctctatgaaa catgggagaa gctctggacc
     2401 aataaaagat tccccaacat cgtcaatcaa tacctcgaga agtggaggga ccgcttctac
     2461 ctcttcgacc agaagtaccc cttcttccag gtcaccaagg aggacatcgc cggcgataaa
     2521 attagcaagg ccaagggcac cagcatcctc ggcaaaaata taaatcgcat catcagcgag
     2581 agcggcaaca agatcgccct cttcagcccc aaggacgagg aaaataaaaa tactctcacc
     2641 gccgccgagc tcgccaggtg gctcatcacc ttccagggct acagcggcgt cagcgacaag
     2701 gtcatcttcg gcaatgaaaa atataccccc agcaagggct ggctcttcga catcggcgcc
     2761 atctacatta aaggcaacac cctcttcgaa accctcctcc tcaactacat cagcccctac
     2821 aacgagtgcg gcaatgtgga aaatatacag aggccctgct gggagaggaa gagcagcgac
     2881 atcattaaaa gctacctcga tgaaaaagac atcaccaaca tcgccagcct ctacaccgtc
     2941 tggagcaagg gcatctacat cgaccccgac ttcaacctca ataaaccctt cagcttcgac
     3001 atcgtcaagc tccccgacat caaccaccgc gacaacttcc tcgagcccat gaccctctgg
     3061 aagtacaacg tcagcggcaa aaatagggac agctacaccc ccaagaagca cctcctcaac
     3121 cagagcctct ggaggagctt cggcctcctg agcataaaag acaccgacct ccagcacagg
     3181 aagcccggcg tcatcgagtg gctcacctac atcgacgata taataggcaa caggctcagc
     3241 aacatcgtcg ccatcagcat gcaggacgac ggcaaccccc agagcaggct gcccaccgac
     3301 gaggtcatcg acagcatctt catcaacgac ctcgtcctca ccgacctcga cgagggcggc
     3361 tgggtcccca ggatcaacga ggtcgtcgag gaaaccaaga agatcatcag caggacatat
     3421 aaaacgtaca ttaatgatat aaaggagatc cgcaacatca gcaacggcag ctacacccag
     3481 cagatcgtcg agagcctcta cttcaagatc gaccagccct tcaggcagtg gctcgccagc
     3541 atccagcccg aggacgacaa ggacagcaag atccaggagt ggcgcgtcct cctcaagaaa
     3601 atagtcaagg ccgaggccga ggtcatcctc caccagggcg gcgccaggga ctacctcggc
     3661 atccagaagg acggccgaat taagaacatc gccaccgcct acaacagctt cgacttctgg
     3721 ctccgccagc agttaaaaag cagggccgac cccaagaaga agaggaaggt gggtcgggcg
     3781 acatgtgcca aggacatcca gaaggccgct gccgaagcgg cgctcgcttt tcaggacgaa
     3841 acctgcgaca ccaccaccac caaccacggc ctcgacatgg aggaaacaat ggtggaggcc
     3901 atctacacgc ctgaacagag cgagggggct ttctacatgg acgaggaaac catgttcggc
     3961 atgcccacac tcctcgacaa catggccgag ggcatgctcc tccccccccc gagcgtccag
     4021 tggaaccaca actacgacgg cgaaggcgac ggcgacgtca gcctctggtc ctactagtct
     4081 tcgtaagtta acctagactt gtccatcttc tggattggcc aacttaatta atgtatgaaa
     4141 taaaaggatg cacacatagt gacatgctaa tcactataat gtgggcatca aagttgtgtg
     4201 ttatgtgtaa ttactagtta tctgaataaa agagaaagag atcatccata tttcttatcc
     4261 taaatgaatg tcacgtgtct ttataattct ttgatgaacc agatgcattt cattaaccaa
     4321 atccatatac atataaatat taatcatata taattaatat caattgggtt agcaaaacaa
     4381 atctagtcta ggtgtgtttt gcgaatgcgg ccgccaccgc ggtggagctc gaattccggt
     4441 ccgggtcacc cagcttgagt attctatagt gtcacctaaa tagcttggcg taatcatggt
     4501 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4561 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     4621 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4681 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4741 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4801 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4861 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4921 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4981 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5041 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     5101 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     5161 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5221 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5281 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5341 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5401 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5461 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5521 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catggagcca cgttgtgtct
     5581 caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg
     5641 tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtct
     5701 tgctcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct
     5761 cgcgataatg tcgggcaatc aggtgcgaca atctatcgat tgtatgggaa gcccgatgcg
     5821 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg
     5881 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt
     5941 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta
     6001 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc
     6061 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc
     6121 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag
     6181 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca
     6241 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg
     6301 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt
     6361 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcag
     6421 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag
     6481 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga
     6541 cgggacggcg gctttgttga ataaatcgaa cttttgctga gttgaaggat cagatcacgc
     6601 atcttcccga caacgcagac cgttccgtgg caaagcaaaa gttcaaaatc accaactggt
     6661 ccacctacaa caaagctctc atcaaccgtg gctccctcac tttctggctg gatgatgggg
     6721 cgattcaggc ctggtatgag tcagcaacac cttcttcacg agccatgaca ttaacctata
     6781 aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc
     6841 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     6901 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg
     6961 cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat
     7021 gcgtaaggag aaaataccgc atcaggaaat tgtaagcgtt aatattttgt taaaattcgc
     7081 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     7141 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     7201 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     7261 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     7321 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     7381 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     7441 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     7501 gtccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     7561 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     7621 gggttttccc agtcacgacg ttgtaaaacg acggccagt
//
