LOCUS       sequence-263429-        2812 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  crRNA backbone for FnCas12a/CmsI nucleases, combine with OsU3 
            promoter module (pFH39).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2812)
  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 2812)
  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: pFH131
FEATURES             Location/Qualifiers
     source          1..2812
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    96..117
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(132..137,138..155,156..162)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    170..186
                     /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     194..210
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             206..529
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    223..279
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(280..296)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(505..524)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(704..720)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(892..1480)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(972..991)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1616..2431)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1616..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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2339..2358
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2423..2442
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     2552..2568
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2552..2568
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     terminator      2569..2615
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2744..2760
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gggtctcagg cataatttct actgttgtag atttgtcttc gcagctggca cgacaggttt
       61 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
      121 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
      181 taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc atgcctgcag
      241 gtcgactcta gaggatcccc gggtaccgag ctcgaattca ctggccgtcg ttttacaacg
      301 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
      361 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
      421 cctgaatggc gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
      481 acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
      541 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
      601 ttacagacaa gctgtgacga agacaatttt tttttcgctt gagaccgagg ctaggtggag
      661 gctcagtgat gataagtctg cgatggtgga tgcatgtgtc atggtcatag ctgtttcctg
      721 tgtgaaattg ttatccgctc agagggcaca atcctattcc gcgctatccg acaatctcca
      781 agacattagg tggagttcag ttcggcgtat ggcatatgtc gctggaaaga acatgtgagc
      841 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
      901 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
      961 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1021 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1081 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1141 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1201 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     1261 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     1321 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     1381 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     1441 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     1501 tacggggtct gacgctctat tcaacaaagc cgccgtcccg tcaagtcagc gtaaatgggt
     1561 agggggcttc aaatcgtccg ctctgccagt gttacaacca attaacaaat tctgattaga
     1621 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     1681 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     1741 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     1801 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     1861 ccggtgagaa tggcaaaagc ttatgcattt ctttccagac ttgttcaaca ggccagccat
     1921 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     1981 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     2041 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     2101 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     2161 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     2221 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     2281 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     2341 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctcg
     2401 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     2461 cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa catcagagat
     2521 tttgagacac aacgtggctt tcccccgccg ctctagaact agtggatcca aataaaacga
     2581 aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcgca ttatacgaga
     2641 cgtccaggtt gggatacctg aaacaaaacc catcgtacgg ccaaggaagt ctccaataac
     2701 tgtgatccac cacaagcgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt
     2761 catgcataat ccgcacgcat ctggaataag gaagtgccat tccgcctgac ct
//
