LOCUS       sequence-252661-        2559 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Endlinker for tRNA-gRNA system when using only five guides.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2559)
  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 2559)
  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: pAK-EL-05
FEATURES             Location/Qualifiers
     source          1..2559
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(469..485)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(469..485)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(482..504)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    493..509
                     /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)."
     promoter        complement(join(517..523,524..541,542..547))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    562..583
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(700..717)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(871..1459)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(951..970)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1637..2446)
                     /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     2360..2379
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2438..2457
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagagaatt cggtctcaag tggcaccgag
      421 tcggtgcttt tttttttttc gcttgagacc aagcttggtg taatcatggt catagctgtt
      481 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
      541 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
      601 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
      661 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
      721 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
      781 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
      841 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
      901 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
      961 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     1021 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     1081 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     1141 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     1201 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     1261 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     1321 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     1381 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     1441 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     1501 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     1561 gatcctttta aattaaaaat gaagttttaa atcaagccca atctgaataa tgttacaacc
     1621 aattaaccaa ttctgattag aaaaactcat cgagcatcaa atgaaactgc aatttattca
     1681 tatcaggatt atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact
     1741 caccgaggca gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc
     1801 caacatcaat acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat
     1861 caccatgagt gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga
     1921 cttgttcaac aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt
     1981 tattcattcg tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat
     2041 tacaaacagg aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt
     2101 cacctgaatc aggatattct tctaatacct ggaatgctgt ttttccgggg atcgcagtgg
     2161 tgagtaacca tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa
     2221 attccgtcag ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt
     2281 tgccatgttt cagaaacaac tctggcgcat cgggcttccc atacaagcga tagattgtcg
     2341 cacctgattg cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt
     2401 tggaatttaa tcgcggcctc gacgtttccc gttgaatatg gctcataaca ccccttgtat
     2461 tactgtttat gtaagcagac agttttattg ttcatgatga tatattttta tcttgtgcaa
     2521 tgtaacatca gagattttga gacacgggcc agagctgca
//
