LOCUS       sequence-252660-        2564 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Endlinker for tRNA-gRNA system when using only four guides.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2564)
  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 2564)
  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-04
FEATURES             Location/Qualifiers
     source          1..2564
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(146..478)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLVSSEKKKSTDSVPLFLRPNSLAVVLQRRDWENPGVTQ
                     LNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLST
                     ICSDAA"
     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(474..490)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(474..490)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(487..509)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    498..514
                     /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(522..528,529..546,547..552))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    567..588
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(705..722)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(876..1464)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(956..975)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1642..2451)
                     /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     2365..2384
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2443..2462
                     /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 cggtctcaga aaaagtggca
      421 ccgagtcggt gctttttttt ttttcgcttg agaccaagct tggtgtaatc atggtcatag
      481 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
      541 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
      601 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
      661 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
      721 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
      781 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
      841 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
      901 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
      961 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     1021 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     1081 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     1141 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     1201 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     1261 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     1321 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     1381 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     1441 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     1501 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     1561 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa gcccaatctg aataatgtta
     1621 caaccaatta accaattctg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt
     1681 attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga
     1741 aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac
     1801 tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga
     1861 gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt
     1921 ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa
     1981 accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg
     2041 acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat
     2101 attttcacct gaatcaggat attcttctaa tacctggaat gctgtttttc cggggatcgc
     2161 agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg
     2221 cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct
     2281 acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca agcgatagat
     2341 tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc
     2401 catgttggaa tttaatcgcg gcctcgacgt ttcccgttga atatggctca taacacccct
     2461 tgtattactg tttatgtaag cagacagttt tattgttcat gatgatatat ttttatcttg
     2521 tgcaatgtaa catcagagat tttgagacac gggccagagc tgca
//
