LOCUS       sequence-254852-        2147 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  guide RNA (gRNA) expression vector used to create a gRNA to a 
            specific sequence; uses T7 promoter for in vitro transcription.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2147)
  AUTHORS   Hwang WY, Fu Y, Reyon D, Maeder ML, Tsai SQ, Sander JD, Peterson RT,
            Yeh JR, Joung JK
  TITLE     Efficient genome editing in zebrafish using a CRISPR-Cas system.
  JOURNAL   Nat Biotechnol. 2013 Jan 29. doi: 10.1038/nbt.2501.
  PUBMED    23360964
REFERENCE   2  (bases 1 to 2147)
  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: "Nat 
            Biotechnol. 2013 Jan 29. doi: 10.1038/nbt.2501."
COMMENT     Sequence Label: DR274
FEATURES             Location/Qualifiers
     source          1..2147
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     9..26
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     10..26
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_RNA        complement(208..283)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(305..323)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(452..468)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(452..468)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      516..547
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        548..650
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             651..1466
                     /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="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(724..743)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1490..1517
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(1529..2116)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1609..1628)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cccacccgtg taaaacgacg gccagtttat ctagtcagct tgattctagc tgatcgtgga
       61 ccggaaggtg agccagtgag ttgattgcag tccagttacg ctggagtctg aggctcgtcc
      121 tgaatgatat gcgaccgccg gagggttgcg tttgagacgg gcgacagatc cagtcgcgct
      181 gctctcgtcg atccaagctt tttaaaagca ccgactcggt gccacttttt caagttgata
      241 acggactagc cttattttaa cttgctattt ctagctctaa aactgagacc ctctctcggt
      301 ctctcctata gtgagtcgta ttagctagcg gtgcgagcgg atcgagcagt gtcgatcact
      361 actggaccgc gagctgtgct gcgacccgtg atcttacggc attatacgta tgatcggtcc
      421 acgatcagct agattatcta gtcagcttga tgtcatagct gtttcctgag gctcaatact
      481 gaccatttaa atcatacctg acctccatag cagaaagtca aaagcctccg accggaggct
      541 tttgacttga tcggcacgta agaggttcca actttcacca taatgaaata agatcactac
      601 cgggcgtatt ttttgagtta tcgagatttt caggagctaa ggaagctaaa atgagccata
      661 ttcaacggga aacgtcttgc ttgaagccgc gattaaattc caacatggat gctgatttat
      721 atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt
      781 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg
      841 atgttacaga tgagatggtc aggctaaact ggctgacgga atttatgcct cttccgacca
      901 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atcccaggga
      961 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc
     1021 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacggcg
     1081 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttggtgcga
     1141 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata
     1201 aactcttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc
     1261 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag
     1321 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac
     1381 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc
     1441 acttgatgct cgatgagttt ttctaatgag gacctaaatg taatcacctg gctcaccttc
     1501 gggtgggcct ttctgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     1561 acaaaaatcg atgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     1621 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     1681 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     1741 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     1801 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     1861 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     1921 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     1981 gtatctgcgc tctgctgaag ccagttacct cggaaaaaga gttggtagct cttgatccgg
     2041 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     2101 aaaaaaagga tctcaagaag atcctttgat tttctaccga agaaagg
//
