LOCUS       sequence-385922-        3136 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for cloning the second CRISPR-Cas9 guide RNA for guide RNAs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3136)
  AUTHORS   Li C, Wang L, Cseke LJ, Vasconcelos F, Huguet-Tapia JC, Gassmann W, 
            Pauwels L, White FF, Dong H, Yang B
  TITLE     Efficient CRISPR-Cas9 based cytosine base editors for 
            phytopathogenic bacteria.
  JOURNAL   Commun Biol. 2023 Jan 17;6(1):56. doi: 10.1038/s42003-023-04451-8.
  PUBMED    36646768
REFERENCE   2  (bases 1 to 3136)
  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: "2023-01-17"; volume: "6(1):56. doi"; pages: " 
            10.1038/s42003-023-04451-8"
COMMENT     Sequence Label: pTL-Bag2T-gRNA2
FEATURES             Location/Qualifiers
     source          1..3136
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(196..201,202..218,219..224)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             272..1462
                     /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(624..643)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(1636..1655)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1637..1655)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1660..1676)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1660..1676)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        1768..1802
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     rep_origin      1985..2573
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2474..2493
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2903..2930)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3022..3108)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
       61 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
      121 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
      181 aattaattct catgtttgac agcttatcat cgattagctt taatgcggta gtttatcaca
      241 gttaaattgc taacgcagtc aggcaccgtg tatgaaatct aacaatgcgc tcatcgtcat
      301 cctcggcacc gtcaccctgg atgctgtagg cataggcttg gttatgccgg tactgccggg
      361 cctcttgcgg gatatcgtcc attccgacag catcgccagt cactatggcg tgctgctagc
      421 gctatatgcg ttgatgcaat ttctatgcgc acccgttctc ggagcactgt ccgaccgctt
      481 tggccgccgc ccagtcctgc tcgcttcgct acttggagcc actatcgact acgcgatcat
      541 ggcgaccaca cccgtcctgt ggattctcta cgccggacgc atcgtggccg gcatcaccgg
      601 cgccacaggt gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc
      661 tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc
      721 cgggggactg ttgggcgcca tctccttaca tgcaccattc cttgcggcgg cggtgctcaa
      781 cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgccg
      841 acccatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac
      901 tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc
      961 agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct
     1021 gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca ctggtcccgc
     1081 caccaaacgt ttcggcgaga agcaggccat tatcgccggc atggcggccg acgcgctggg
     1141 ctacgtcttg ctggcgttcg cgacgcgagg ctggatggcc ttccccatta tgattcttct
     1201 cgcttccggc ggcatcggga tgcccgcgtt gcaggccatg ctgtccaggc aggtagatga
     1261 cgaccatcag ggacagcttc aaggatcgct cgcggctctt accagcctaa cttcgatcat
     1321 tggaccgctg atcgtcacgg cgatttatgc cgcctcggcg agcacatgga acgggttggc
     1381 atggattgta ggcgccgccc tataccttgt ctgcctcccc gcgttgcgtc gcggtgcatg
     1441 gagccgggcc acctcgacct gaatggaagc cggcggcacc tcgctaacgg attcaccact
     1501 ccaagaattg gagccaatca attcttgcgg agaactgtga atgcgcaaac caacccttga
     1561 tcggggaaga acagtatgtc gagctatttt ttgacttact ggggatcaag cctgattggg
     1621 agaaaataaa atatccccta tagtgagtcg tattacatgg tcatagctgt ttcctggcag
     1681 ctctggcccg tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca
     1741 tgcctcctct agaatgggtc tctctgattg acagctagct cagtcctagg tataatgcta
     1801 gcggagacga agcttgaaga ccgtcgacct gcagactggc tgtgtataag ggagcctgac
     1861 atttatattc ccccgtctcg gtttggagac catgctcgag ccacccatga ccaaaatccc
     1921 ttaacgtgag ttacgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     1981 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2041 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2101 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     2161 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2221 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2281 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2341 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2401 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     2461 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     2521 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     2581 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     2641 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     2701 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     2761 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     2821 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatacgcgt accgctagcc
     2881 aggaagagtt tgtagaaacg caaaaaggcc atccgtcagg atggccttct gcttagtttg
     2941 atgcctggca gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt gcttcacaac
     3001 gttcaaatcc gctcccggcg gatttgtcct actcaggaga gcgttcaccg acaaacaaca
     3061 gataaaacga aaggcccagt cttccgactg agcctttcgt tttatttgat gcctggcagt
     3121 tccctactct cgcgtt
//
