LOCUS       sequence-271523-        2812 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  crRNA backbone for FnCas12a/CmsI nucleases, combine with OsU6-2 
            promoter module (pFH37).
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: pFH130
FEATURES             Location/Qualifiers
     source          1..2812
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     12..28
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    176..197
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(212..217,218..235,236..242)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    250..266
                     /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     274..290
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             286..609
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    303..359
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(360..376)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(585..604)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(784..800)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(972..1560)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1052..1071)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1696..2511)
                     /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(1696..2511)
                     /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     2419..2438
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2503..2522
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     2632..2648
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2632..2648
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     terminator      2649..2695
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gtcacgacgt tgtaaaacga cggccagtca tgcataatcc gcacgcatct ggaataagga
       61 agtgccattc cgcctgacct gggtctcagt tgtaatttct actgttgtag atttgtcttc
      121 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
      181 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
      241 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
      301 ccaagcttgc atgcctgcag gtcgactcta gaggatcccc gggtaccgag ctcgaattca
      361 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
      421 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
      481 ccttcccaac agttgcgcag cctgaatggc gaatggcgcc tgatgcggta ttttctcctt
      541 acgcatctgt gcggtatttc acaccgcata tggtgcactc tcagtacaat ctgctctgat
      601 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
      661 tgtctgctcc cggcatccgc ttacagacaa gctgtgacga agacaatttt tttttcgctt
      721 gagaccgagg ctaggtggag gctcagtgat gataagtctg cgatggtgga tgcatgtgtc
      781 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc agagggcaca atcctattcc
      841 gcgctatccg acaatctcca agacattagg tggagttcag ttcggcgtat ggcatatgtc
      901 gctggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
      961 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     1021 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     1081 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     1141 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     1201 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     1261 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     1321 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     1381 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     1441 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     1501 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     1561 gaagatcctt tgatcttttc tacggggtct gacgctctat tcaacaaagc cgccgtcccg
     1621 tcaagtcagc gtaaatgggt agggggcttc aaatcgtccg ctctgccagt gttacaacca
     1681 attaacaaat tctgattaga aaaactcatc gagcatcaaa tgaaactgca atttattcat
     1741 atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc
     1801 accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc
     1861 aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc
     1921 accatgagtg acgactgaat ccggtgagaa tggcaaaagc ttatgcattt ctttccagac
     1981 ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt
     2041 attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt
     2101 acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc
     2161 acctgaatca ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt
     2221 gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa
     2281 ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt
     2341 gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc
     2401 acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt
     2461 ggaatttaat cgcggcctcg agcaagacgt ttcccgttga atatggctca taacacccct
     2521 tgtattactg tttatgtaag cagacagttt tattgttcat gatgatatat ttttatcttg
     2581 tgcaatgtaa catcagagat tttgagacac aacgtggctt tcccccgccg ctctagaact
     2641 agtggatcca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     2701 gtttgtcgca ttatacgaga cgtccaggtt gggatacctg aaacaaaacc catcgtacgg
     2761 ccaaggaagt ctccaataac tgtgatccac cacaagcgcc agggttttcc ca
//
