LOCUS       sequence-252817-        2594 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  classic sgRNA backbone for tRNA-gRNA array, Position 5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2594)
  AUTHORS   Hahn F, Korolev A, Sanjurjo Loures L, Nekrasov V
  TITLE     A modular cloning toolkit for genome editing in plants.
  JOURNAL   BMC Plant Biol. 2020 Apr 23;20(1):179. doi: 
            10.1186/s12870-020-02388-2.
  PUBMED    32326901
REFERENCE   2  (bases 1 to 2594)
  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: "BMC Plant 
            Biol."; date: "2020-04-23"; volume: "20(1):179. doi"; pages: " 
            10.1186/s12870-020-02388-2"
COMMENT     Sequence Label: pAK011
FEATURES             Location/Qualifiers
     source          1..2594
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     protein_bind    237..258
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(273..278,279..296,297..303)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    311..327
                     /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)."
     primer_bind     316..338
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     335..351
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     335..351
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             347..670
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    364..420
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(421..438)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(421..437)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(430..452)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(646..665)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(906..1494)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(986..1005)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1672..2481)
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2395..2414
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2473..2492
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gacgtcattg
       61 tcgatggtct cagaaaaagt ggcaccgagt cggtgcaaca aagcaccagt ggtctagtgg
      121 tagaatagta ccctgccacg gtacagaccc gggttcgatt cccggctggt gcattgtctt
      181 cgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
      241 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
      301 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
      361 gccaagcttg catgcctgca ggtcgactct agaggatccc cgggtaccga gctcgaattc
      421 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
      481 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
      541 cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt attttctcct
      601 tacgcatctg tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga
      661 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
      721 ttgtctgctc ccggcatccg cttacagaca agctgtgacg aagacaagtt ttagagctag
      781 aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtgtg agaccatcat
      841 tgtcacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
      901 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
      961 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     1021 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     1081 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     1141 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     1201 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     1261 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     1321 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     1381 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     1441 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     1501 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     1561 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     1621 tttaaatcaa gcccaatctg aataatgtta caaccaatta accaattctg attagaaaaa
     1681 ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa taccatattt
     1741 ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc ataggatggc
     1801 aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac ctattaattt
     1861 cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga ctgaatccgg
     1921 tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc agccattacg
     1981 ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt gcgcctgagc
     2041 gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg aatgcaaccg
     2101 gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat attcttctaa
     2161 tacctggaat gctgtttttc cggggatcgc agtggtgagt aaccatgcat catcaggagt
     2221 acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt ttagtctgac
     2281 catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa acaactctgg
     2341 cgcatcgggc ttcccataca agcgatagat tgtcgcacct gattgcccga cattatcgcg
     2401 agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg gcctcgacgt
     2461 ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag cagacagttt
     2521 tattgttcat gatgatatat ttttatcttg tgcaatgtaa catcagagat tttgagacac
     2581 gggccagagc tgca
//
