LOCUS       sequence-252874-        2544 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  classic sgRNA backbone for tRNA-gRNA array, Position 1, combine with
            OsU6-2 promoter module (pFH36).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2544)
  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 2544)
  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: pFH73
FEATURES             Location/Qualifiers
     source          1..2544
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     protein_bind    218..239
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(254..259,260..277,278..284)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    292..308
                     /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     297..319
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     316..332
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     316..332
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             328..651
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    345..401
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(402..419)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(402..418)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(411..433)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(627..646)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(856..1444)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(936..955)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1622..2431)
                     /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     2345..2364
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2423..2442
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gacgtcattg
       61 tcgatggtct catgttgaac aaagcaccag tggtctagtg gtagaatagt accctgccac
      121 ggtacagacc cgggttcgat tcccggctgg tgcattgtct tcgcagctgg cacgacaggt
      181 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
      241 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
      301 gataacaatt tcacacagga aacagctatg accatgatta cgccaagctt gcatgcctgc
      361 aggtcgactc tagaggatcc ccgggtaccg agctcgaatt cactggccgt cgttttacaa
      421 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
      481 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
      541 agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
      601 tcacaccgca tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
      661 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
      721 gcttacagac aagctgtgac gaagacaagt tttagagcta gaaatagcaa gttaaaattg
      781 agaccatcat tgtcacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
      841 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
      901 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
      961 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     1021 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     1081 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     1141 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     1201 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     1261 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     1321 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     1381 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     1441 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     1501 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     1561 aaaatgaagt tttaaatcaa gcccaatctg aataatgtta caaccaatta accaattctg
     1621 attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa
     1681 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc
     1741 ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac
     1801 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga
     1861 ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc
     1921 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt
     1981 gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg
     2041 aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat
     2101 attcttctaa tacctggaat gctgtttttc cggggatcgc agtggtgagt aaccatgcat
     2161 catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt
     2221 ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa
     2281 acaactctgg cgcatcgggc ttcccataca agcgatagat tgtcgcacct gattgcccga
     2341 cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg
     2401 gcctcgacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     2461 cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa catcagagat
     2521 tttgagacac gggccagagc tgca
//
