LOCUS       sequence-254112-        6289 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CasB constitutive expression cassette for Zea mays.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6289)
  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 6289)
  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: PV244
FEATURES             Location/Qualifiers
     source          1..6289
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..43
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     35..52
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     36..52
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     59..78
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        59..77
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        112..2107
                     /label=Ubi promoter
                     /note="maize polyubiquitin gene promoter"
     CDS             2729..2749
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        complement(3138..3156)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3139..3156)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3174..3190)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3174..3190)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3187..3209)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3198..3214
                     /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)."
     protein_bind    3274..3295
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3412..3429)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3583..4171)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3663..3682)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4345..4364)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4356..5171
                     /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             4356..5171
                     /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(4429..4448)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5465..5483
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5521..5543)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5643..5662
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(5729..6184)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5866..5887)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6078..6097
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaattgta
       61 atacgactca ctatagggcg aattgggtta cccggaccga agcttgcatg cctgcagtgc
      121 agcgtgaccc ggtcgtgccc ctctctagag ataatgagca ttgcatgtct aagttataaa
      181 aaattaccac atattttttt tgtcacactt gtttgaagtg cagtttatct atctttatac
      241 atatatttaa actttactct acgaataata taatctatag tactacaata atatcagtgt
      301 tttagagaat catataaatg aacagttaga catggtctaa aggacaattg agtattttga
      361 caacaggact ctacagtttt atctttttag tgtgcatgtg ttctcctttt tttttgcaaa
      421 tagcttcacc tatataatac ttcatccatt ttattagtac atccatttag ggtttagggt
      481 taatggtttt tatagactaa tttttttagt acatctattt tattctattt tagcctctaa
      541 attaagaaaa ctaaaactct attttagttt ttttatttaa taatttagat ataaaataga
      601 ataaaataaa gtgactaaaa attaaacaaa taccctttaa gaaattaaaa aaactaagga
      661 aacatttttc ttgtttcgag tagataatgc cagcctgtta aacgccgtcg acgagtctaa
      721 cggacaccaa ccagcgaacc agcagcgtcg cgtcgggcca agcgaagcag acggcacggc
      781 atctctgtcg ctgcctctgg acccctctcg agagttccgc tccaccgttg gacttgctcc
      841 gctgtcggca tccagaaatt gcgtggcgga gcggcagacg tgagccggca cggcaggcgg
      901 cctcctcctc ctctcacggc accggcagct acgggggatt cctttcccac cgctccttcg
      961 ctttcccttc ctcgcccgcc gtaataaata gacaccccct ccacaccctc tttccccaac
     1021 ctcgtgttgt tcggagcgca cacacacaca accagatctc ccccaaatcc acccgtcggc
     1081 acctccgctt caaggtacgc cgctcgtcct cccccccccc cctctctacc ttctctagat
     1141 cggcgttccg gtccatgcat ggttagggcc cggtagttct acttctgttc atgtttgtgt
     1201 tagatccgtg tttgtgttag atccgtgctg ctagcgttcg tacacggatg cgacctgtac
     1261 gtcagacacg ttctgattgc taacttgcca gtgtttctct ttggggaatc ctgggatggc
     1321 tctagccgtt ccgcagacgg gatcgatttc atgatttttt ttgtttcgtt gcatagggtt
     1381 tggtttgccc ttttccttta tttcaatata tgccgtgcac ttgtttgtcg ggtcatcttt
     1441 tcatgctttt ttttgtcttg gttgtgatga tgtggtctgg ttgggcggtc gttctagatc
     1501 ggagtagaat tctgtttcaa actacctggt ggatttatta attttggatc tgtatgtgtg
     1561 tgccatacat attcatagtt acgaattgaa gatgatggat ggaaatatcg atctaggata
     1621 ggtatacatg ttgatgcggg ttttactgat gcatatacag agatgctttt tgttcgcttg
     1681 gttgtgatga tgtggtgtgg ttgggcggtc gttcattcgt tctagatcgg agtagaatac
     1741 tgtttcaaac tacctggtgt atttattaat tttggaactg tatgtgtgtg tcatacatct
     1801 tcatagttac gagtttaaga tggatggaaa tatcgatcta ggataggtat acatgttgat
     1861 gtgggtttta ctgatgcata tacatgatgg catatgcagc atctattcat atgctctaac
     1921 cttgagtacc tatctattat aataaacaag tatgttttat aattattttg atcttgatat
     1981 acttggatga tggcatatgc agcagctata tgtggatttt tttagccctg ccttcatacg
     2041 ctatttattt gcttggtact gtttcttttg tcgatgctca ccctgttgtt tggtgttact
     2101 tctgcaggtc gactctagag gatccatgag ccagcatact aaacagaccg tctggaccgt
     2161 caccaagaag atcatcttca agctcagcaa ccaaaaggag gaaacctacg acaaggccac
     2221 cctcgccaag gtcagaaaga ccctcggcaa gcccctcagc gaggccaccg agatctggcc
     2281 catcctcttc gagaacctcc ccgaggagtt cctcagcagc taccagcaac caagctacga
     2341 ggagctcgcc atctacaccg ccctccagct ctacgccctc caccagcagg gcgtcagcac
     2401 ctccgtcatg ctggacgaca gcaagcccta tgaaaatata ggcagcgccc tcgggaagct
     2461 gaggaaggag gacgacacca ccagcatcga caggaggttc aacaccatga tcaccagcag
     2521 caccttcgag gagctcacct accacctcag gcacctcatc agcctcctca aggtcgagtc
     2581 ccccgtcatc caggtcgact acagcggcct cgccaacgac ctctactggt tcctcaataa
     2641 tagccaggag aacgtcaggc tcaactgggc ccgcgcctac tacaagcaaa ttaaaggcga
     2701 gaagaacaac gacaacagcc gcgccgaccc caagaagaag aggaaggtct agtcttcgta
     2761 agttaaccta gacttgtcca tcttctggat tggccaactt aattaatgta tgaaataaaa
     2821 ggatgcacac atagtgacat gctaatcact ataatgtggg catcaaagtt gtgtgttatg
     2881 tgtaattact agttatctga ataaaagaga aagagatcat ccatatttct tatcctaaat
     2941 gaatgtcacg tgtctttata attctttgat gaaccagatg catttcatta accaaatcca
     3001 tatacatata aatattaatc atatataatt aatatcaatt gggttagcaa aacaaatcta
     3061 gtctaggtgt gttttgcgaa tgcggccgcc accgcggtgg agctcgaatt ccggtccggg
     3121 tcacccagct tgagtattct atagtgtcac ctaaatagct tggcgtaatc atggtcatag
     3181 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     3241 ataaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     3301 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     3361 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     3421 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     3481 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     3541 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3601 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3661 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     3721 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3781 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3841 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3901 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3961 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     4021 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     4081 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     4141 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     4201 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatggagc cacgttgtgt
     4261 ctcaaaatct ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac
     4321 tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt
     4381 cttgctcgag gccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg
     4441 ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg attgtatggg aagcccgatg
     4501 cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga
     4561 tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc
     4621 gtactcctga tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca gcattccagg
     4681 tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc
     4741 gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc
     4801 tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg
     4861 agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt ttgccattct
     4921 caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg
     4981 ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc
     5041 ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc
     5101 agaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg
     5161 agtttttcta atcagaattg gttaattggt tgtaacactg gcagagcatt acgctgactt
     5221 gacgggacgg cggctttgtt gaataaatcg aacttttgct gagttgaagg atcagatcac
     5281 gcatcttccc gacaacgcag accgttccgt ggcaaagcaa aagttcaaaa tcaccaactg
     5341 gtccacctac aacaaagctc tcatcaaccg tggctccctc actttctggc tggatgatgg
     5401 ggcgattcag gcctggtatg agtcagcaac accttcttca cgagccatga cattaaccta
     5461 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     5521 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     5581 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta
     5641 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     5701 atgcgtaagg agaaaatacc gcatcaggaa attgtaagcg ttaatatttt gttaaaattc
     5761 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
     5821 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
     5881 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
     5941 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa
     6001 gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg
     6061 aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt
     6121 gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc
     6181 gcgtccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     6241 cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaag
//
