LOCUS       sequence-391280-        3618 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Golden Gate entry vector; 3rd gRNA under TaU3 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3618)
  AUTHORS   Lee K, Kang M, Ji Q, Grosic S, Wang K
  TITLE     New T-DNA binary vectors with NptII selection and RUBY reporter for 
            efficient maize transformation and targeted mutagenesis.
  JOURNAL   Plant Physiol. 2023 Apr 18:kiad231. doi: 10.1093/plphys/kiad231.
  PUBMED    37070560
REFERENCE   2  (bases 1 to 3618)
  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: "Plant 
            Physiol. 2023 Apr 18:kiad231. doi: 10.1093/plphys/kiad231."
COMMENT     Sequence Label: pCBLsgRNA3
FEATURES             Location/Qualifiers
     source          1..3618
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(189..216)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(308..394)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        join(618..623,624..640,641..646)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             694..1884
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(1046..1065)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(2058..2077)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2059..2077)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2082..2098)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2082..2098)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      2889..3477
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3378..3397
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
       61 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
      121 aaagcgggca gtgagcgcaa cgcaattaat acgcgtaccg ctagccagga agagtttgta
      181 gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt agtttgatgc ctggcagttt
      241 atggcgggcg tcctgcccgc caccctccgg gccgttgctt cacaacgttc aaatccgctc
      301 ccggcggatt tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg
      361 cccagtcttc cgactgagcc tttcgtttta tttgatgcct ggcagttccc tactctcgcg
      421 ttctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
      481 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
      541 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
      601 tgaattaatt ctcatgtttg acagcttatc atcgattagc tttaatgcgg tagtttatca
      661 cagttaaatt gctaacgcag tcaggcaccg tgtatgaaat ctaacaatgc gctcatcgtc
      721 atcctcggca ccgtcaccct ggatgctgta ggcataggct tggttatgcc ggtactgccg
      781 ggcctcttgc gggatatcgt ccattccgac agcatcgcca gtcactatgg cgtgctgcta
      841 gcgctatatg cgttgatgca atttctatgc gcacccgttc tcggagcact gtccgaccgc
      901 tttggccgcc gcccagtcct gctcgcttcg ctacttggag ccactatcga ctacgcgatc
      961 atggcgacca cacccgtcct gtggattctc tacgccggac gcatcgtggc cggcatcacc
     1021 ggcgccacag gtgcggttgc tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg
     1081 gctcgccact tcgggctcat gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg
     1141 gccgggggac tgttgggcgc catctcctta catgcaccat tccttgcggc ggcggtgctc
     1201 aacggcctca acctactact gggctgcttc ctaatgcagg agtcgcataa gggagagcgc
     1261 cgacccatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg
     1321 actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg
     1381 gcagcgctct gggtcatttt cggcgaggac cgctttcgct ggagcgcgac gatgatcggc
     1441 ctgtcgcttg cggtattcgg aatcttgcac gccctcgctc aagccttcgt cactggtccc
     1501 gccaccaaac gtttcggcga gaagcaggcc attatcgccg gcatggcggc cgacgcgctg
     1561 ggctacgtct tgctggcgtt cgcgacgcga ggctggatgg ccttccccat tatgattctt
     1621 ctcgcttccg gcggcatcgg gatgcccgcg ttgcaggcca tgctgtccag gcaggtagat
     1681 gacgaccatc agggacagct tcaaggatcg ctcgcggctc ttaccagcct aacttcgatc
     1741 attggaccgc tgatcgtcac ggcgatttat gccgcctcgg cgagcacatg gaacgggttg
     1801 gcatggattg taggcgccgc cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca
     1861 tggagccggg ccacctcgac ctgaatggaa gccggcggca cctcgctaac ggattcacca
     1921 ctccaagaat tggagccaat caattcttgc ggagaactgt gaatgcgcaa accaaccctt
     1981 gatcggggaa gaacagtatg tcgagctatt ttttgactta ctggggatca agcctgattg
     2041 ggagaaaata aaatatcccc tatagtgagt cgtattacat ggtcatagct gtttcctggc
     2101 agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat
     2161 catgcctcct ctagaggtct cgggacggcg gcagggagag ttttaacatt gactagcgtg
     2221 ctgataattt gtgagaaata ataattgaca agtagatact gacatttgag aagagcttct
     2281 gaactgttat tagtaacaaa aatggaaagc tgatgcacgg aaaaaggaaa gaaaaagcca
     2341 tacttttttt taggtaggaa aagaaaaagc catacgagac tgatgtctct cagatgggcc
     2401 gggatctgtc tatctagcag gcagcagccc taccaacctc acgggccagc aattacgagt
     2461 ccttctaaaa cgtcccgccg agggcgcgtg gccgtgctgt gcagcagcac gtctaacatt
     2521 agtcccacct cgccagttta cagggagcag aaccagctta taagcggagg cgcggcacca
     2581 agaagcgaga gacgagatct agtctgagtc gaccgtctct gtttcagagc tatgctggaa
     2641 acagcatagc aagttgaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt
     2701 cggtgctttt tttgtccctt cgaagggcaa ttctgcagat atccatcaca ctggcggccg
     2761 ctcgaggtcg agggtatcga taagcttcca gggagaccct cgagccaccc atgaccaaaa
     2821 tcccttaacg tgagttacgc gtcgttccac tgagcgtcag accccgtaga aaagatcaaa
     2881 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     2941 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3001 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     3061 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3121 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3181 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     3241 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     3301 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     3361 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     3421 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     3481 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     3541 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     3601 accgctcgcc gcagccga
//
