LOCUS       sequence-252902-        2554 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  improved 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 2554)
  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 2554)
  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: pFH50
FEATURES             Location/Qualifiers
     source          1..2554
                     /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(866..1454)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(946..965)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1632..2441)
                     /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     2355..2374
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2433..2452
                     /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 ttcagagcta tgctggaaac agcatagcaa
      781 gttgaaattg agaccatcat tgtcacatgt gagcaaaagg ccagcaaaag gccaggaacc
      841 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
      901 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
      961 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     1021 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     1081 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     1141 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     1201 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     1261 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     1321 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     1381 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     1441 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     1501 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     1561 ttttaaatta aaaatgaagt tttaaatcaa gcccaatctg aataatgtta caaccaatta
     1621 accaattctg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca
     1681 ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg
     1741 aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca
     1801 tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca
     1861 tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt
     1921 tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc
     1981 attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa
     2041 acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct
     2101 gaatcaggat attcttctaa tacctggaat gctgtttttc cggggatcgc agtggtgagt
     2161 aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc
     2221 gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca
     2281 tgtttcagaa acaactctgg cgcatcgggc ttcccataca agcgatagat tgtcgcacct
     2341 gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa
     2401 tttaatcgcg gcctcgacgt ttcccgttga atatggctca taacacccct tgtattactg
     2461 tttatgtaag cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa
     2521 catcagagat tttgagacac gggccagagc tgca
//
