LOCUS       sequence-258473-        6630 bp    DNA     circular SYN 15-APR-2025
DEFINITION  CasE 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 6630)
  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 6630)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: PV503
FEATURES             Location/Qualifiers
     source          1..6630
                     /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             2722..2742
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        complement(3434..3452)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3435..3452)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3470..3486)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3470..3486)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3483..3505)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3494..3510
                     /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(3518..3524,3525..3542,3543..3548))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3563..3584
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3701..3718)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3872..4460)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3952..3971)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4634..4653)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4645..5460
                     /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             4645..5460
                     /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(4718..4737)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5754..5772
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5810..5832)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5932..5951
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(6018..6473)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6155..6176)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6367..6386
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6599..6621
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6613..6630
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6614..6630
                     /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 ctcgtcctnc cccccccccc 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 tccatgtaca tcagcagggt cgagatcggc
     2101 cgatataata ggcgcaaggt cagggacctc acccacgtcg gcgcctacca cgcctgggtc
     2161 gaggagagct tccccagcga gctcgagcaa tcaatcagga ccaggaagct ctggcgcatc
     2221 gaccgcatcc agggcaagga ctacctcatc atcgtcagca aggagaagcc caacctccag
     2281 aagctcgaaa aatatggcgt cgtcggcagc gcccagacca aggactacca gcacttcctc
     2341 gacagcatca agaccggctt caggatgaac ttcaggatcg tcctcaaccc cgtcatcagc
     2401 atcaccagcc ccgacaaatc aaaacgcggc gtcgtcaagc cccacgtcac cattaaacac
     2461 cagatggact tcctcgtcaa gcagagcgaa aaaaacggct tcagcctcgt cggcgagagc
     2521 ttctccatcg tcgagagggg ctacgaggtg ttccagaagc ccgagaagcc catcaggctc
     2581 agcaaggtcg tctacgaggg caccctcacc gtcagcgacg acacgctctt caagaagatg
     2641 ctcaccgagg gcgtcggcaa gaagaaagca tatggcttcg gcctcatgac cgtcgtcccc
     2701 ctcgaggaga gcagggccga ccccaagaag aagaggaagg tgggtcgggc gacatgtgcc
     2761 aaggacatcc agaaggccgc tgccgaagcg gcgctcgctt ttcaggacga aacctgcgac
     2821 accaccacca ccaaccacgg cctcgacatg gaggaaacaa tggtggaggc catctacacg
     2881 cctgaacaga gcgagggggc tttctacatg gacgaggaaa ccatgttcgg catgcccaca
     2941 ctcctcgaca acatggccga gggcatgctc ctcccccccc cgagcgtcca gtggaaccac
     3001 aactacgacg gcgaaggcga cggcgacgtc agcctctggt cctactagtc ttcgtaagtt
     3061 aacctagact tgtccatctt ctggattggc caacttaatt aatgtatgaa ataaaaggat
     3121 gcacacatag tgacatgcta atcactataa tgtgggcatc aaagttgtgt gttatgtgta
     3181 attactagtt atctgaataa aagagaaaga gatcatccat atttcttatc ctaaatgaat
     3241 gtcacgtgtc tttataattc tttgatgaac cagatgcatt tcattaacca aatccatata
     3301 catataaata ttaatcatat ataattaata tcaattgggt tagcaaaaca aatctagtct
     3361 aggtgtgttt tgcgaatgcg gccgccaccg cggtggagct cgaattccgg tccgggtcac
     3421 ccagcttgag tattctatag tgtcacctaa atagcttggc gtaatcatgg tcatagctgt
     3481 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     3541 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     3601 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     3661 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     3721 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     3781 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     3841 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     3901 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     3961 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4021 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4081 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4141 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4201 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4261 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     4321 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4381 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4441 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4501 ggaacgaaaa ctcacgttaa gggattttgg tcatggagcc acgttgtgtc tcaaaatctc
     4561 tgatgttaca ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac
     4621 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctcgagg
     4681 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     4741 gtcgggcaat caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg
     4801 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     4861 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     4921 gatgcatggt tactcaccac tgcgatcccc ggaaaaacag cattccaggt attagaagaa
     4981 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat
     5041 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     5101 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     5161 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     5221 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata
     5281 ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta
     5341 tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca gaaatatggt
     5401 attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa
     5461 tcagaattgg ttaattggtt gtaacactgg cagagcatta cgctgacttg acgggacggc
     5521 ggctttgttg aataaatcga acttttgctg agttgaagga tcagatcacg catcttcccg
     5581 acaacgcaga ccgttccgtg gcaaagcaaa agttcaaaat caccaactgg tccacctaca
     5641 acaaagctct catcaaccgt ggctccctca ctttctggct ggatgatggg gcgattcagg
     5701 cctggtatga gtcagcaaca ccttcttcac gagccatgac attaacctat aaaaataggc
     5761 gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
     5821 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
     5881 gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag
     5941 agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga tgcgtaagga
     6001 gaaaataccg catcaggaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     6061 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     6121 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     6181 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     6241 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     6301 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     6361 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     6421 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtccattcg
     6481 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     6541 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     6601 cagtcacgac gttgtaaaac gacggccagt
//
