LOCUS       sequence-391369-        3395 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Golden Gate entry vector; 4th gRNA under ZmU6-C1 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3395)
  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 3395)
  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: pCBLsgRNA4
FEATURES             Location/Qualifiers
     source          1..3395
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..19)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(24..40)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(24..40)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      608..1196
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1097..1116
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1526..1553)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(1645..1731)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        join(1955..1960,1961..1977,1978..1983)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             2031..3221
                     /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(2383..2402)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(join(3395,1..19))
                     /label=T7
                     /note="T7 promoter, forward primer"
ORIGIN
        1 cctatagtga gtcgtattac atggtcatag ctgtttcctg gcagctctgg cccgtgtctc
       61 aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgcctc ctctagaggt
      121 ctcgccagca cgtgagcttg cgatgtccac tagggagctc catccactga tccaccccca
      181 cgcggcgtgg cgtcgtcatt aacggcttgt ggggaaggga acgagcaact aaccgataat
      241 tagtaccaga ccggccagtg aacgatgcca aaaccggctt ataagctcag ctgcgacaac
      301 cgttttggag acgagatcta gtctgagtcg accgtctctg tttcagagct atgctggaaa
      361 cagcatagca agttgaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
      421 ggtgcttttt ttgtcccttc gaagggcaat tctgcagata tccatcacac tggcggccgc
      481 tcgaggtcga gggtatcgat aagctttgtt ggagaccctc gagccaccca tgaccaaaat
      541 cccttaacgt gagttacgcg tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
      601 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
      661 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
      721 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
      781 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
      841 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
      901 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
      961 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     1021 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     1081 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     1141 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     1201 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     1261 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     1321 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     1381 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     1441 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatacg cgtaccgcta
     1501 gccaggaaga gtttgtagaa acgcaaaaag gccatccgtc aggatggcct tctgcttagt
     1561 ttgatgcctg gcagtttatg gcgggcgtcc tgcccgccac cctccgggcc gttgcttcac
     1621 aacgttcaaa tccgctcccg gcggatttgt cctactcagg agagcgttca ccgacaaaca
     1681 acagataaaa cgaaaggccc agtcttccga ctgagccttt cgttttattt gatgcctggc
     1741 agttccctac tctcgcgttc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     1801 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     1861 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     1921 taacgcagga aagaacatga attaattctc atgtttgaca gcttatcatc gattagcttt
     1981 aatgcggtag tttatcacag ttaaattgct aacgcagtca ggcaccgtgt atgaaatcta
     2041 acaatgcgct catcgtcatc ctcggcaccg tcaccctgga tgctgtaggc ataggcttgg
     2101 ttatgccggt actgccgggc ctcttgcggg atatcgtcca ttccgacagc atcgccagtc
     2161 actatggcgt gctgctagcg ctatatgcgt tgatgcaatt tctatgcgca cccgttctcg
     2221 gagcactgtc cgaccgcttt ggccgccgcc cagtcctgct cgcttcgcta cttggagcca
     2281 ctatcgacta cgcgatcatg gcgaccacac ccgtcctgtg gattctctac gccggacgca
     2341 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca
     2401 ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta
     2461 tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttacat gcaccattcc
     2521 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt
     2581 cgcataaggg agagcgccga cccatgccct tgagagcctt caacccagtc agctccttcc
     2641 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac
     2701 tcgtaggaca ggtgccggca gcgctctggg tcattttcgg cgaggaccgc tttcgctgga
     2761 gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag
     2821 ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt atcgccggca
     2881 tggcggccga cgcgctgggc tacgtcttgc tggcgttcgc gacgcgaggc tggatggcct
     2941 tccccattat gattcttctc gcttccggcg gcatcgggat gcccgcgttg caggccatgc
     3001 tgtccaggca ggtagatgac gaccatcagg gacagcttca aggatcgctc gcggctctta
     3061 ccagcctaac ttcgatcatt ggaccgctga tcgtcacggc gatttatgcc gcctcggcga
     3121 gcacatggaa cgggttggca tggattgtag gcgccgccct ataccttgtc tgcctccccg
     3181 cgttgcgtcg cggtgcatgg agccgggcca cctcgacctg aatggaagcc ggcggcacct
     3241 cgctaacgga ttcaccactc caagaattgg agccaatcaa ttcttgcgga gaactgtgaa
     3301 tgcgcaaacc aacccttgat cggggaagaa cagtatgtcg agctattttt tgacttactg
     3361 gggatcaagc ctgattggga gaaaataaaa tatcc
//
