If you need ZKO Alleles,please contact luyuanpan@ihb.ac.cn
ZKO number ZKO1000
Recent progress of this line F2 individuals have been indentified
Gene name spred2a
Ensembl ID ENSDARG00000052435
Zebrafish strain AB
Cas9 target site 1 sequence sense strand
GGCGGAGGTCTCAGTCGGGTggg
Cas9 target site 2 sequence
Position in the gene Exon 2
Restriction enzyme (RE) n/a
PCR primer (forward) TTTAGAGCAGATTTCCTCCTTT
PCR primer (reverse) AATGAGCCTAGATTGTGTGTGTA
Length of expected PCR product (bp) 345
Length of expected RE digestion product (bp)
Plasmids used for making Cas9 mRNA pXT7-hCAS9
Plasmids used for making gRNA pMD19T-gRNA
Primers used for making gRNA Pr-ZKO
Dosage of injected Cas9 mRNA (ng/ul) 400
Dosage of injected gRNA (ng/ul) 50
Survival rate of F0 embryo @ 1dpf (%) 80
Method to estimate mutation efficiency sequencing of PCR product
Stage of the embryos collected for mutation detection http://www.zfish.cn/upload/file/ZKOPDF/ZKO1000.pdf
Mutation efficiency in F0 embryos (%) 40
Picture from F0 embryos
  (gel picture or sequencing picture)
Mutation type Indel
Germline transmission efficiency in adult F0 27%(3/11)
Wild-type (WT) sequence CTGGCTCAGGAGGGCGGAGGTCTCAGTCGGGTGGGCGTCTGTAAGGTGGTCCCGGCTGAT
Germline mutations in selected F0
 (By screening F1 embryos)
F0-1 GGCTGGCTCAGGAGGGCGGAGGTCTCAGTCGGTGCCACTTTTTCAAGTTGATAACGGACTAGCCTTATTTTAACTTGCTATTTCTAGCTCTAAAAAAATTTCTTTTAGCTCTAAAACACCCGACTGTAAGGTGGTCCCGGCTGAT (-8,+91)
F0-2 GGCTGGCTCAGGAGGGCGGAGGTCTCAGT-----GGGCGTCTGTAAGGTGGTCCCGGCT (-5)
Mosaicism in selected F0
 (By screening F1 adults)
F0-1 GGCTGGCTCAGGAGGGCGGAGGTCTCAGTCGGTGCCACTTTTTCAAGTTGATAACGGACTAGCCTTATTTTAACTTGCTATTTCTAGCTCTAAAAAAATTTCTTTTAGCTCTAAAACACCCGACTGTAAGGTGGTCCCGGCTGAT (-8,+91)
F0-2 GGCTGGCTCAGGAGGGCGGAGGTCTCAGT-----GGGCGTCTGTAAGGTGGTCCCGGCT (-5)
Allele name and predicted product 1000a
1000b
GGCTGGCTCAGGAGGGCGGAGGTCTCAGTCGGTGCCACTTTTTCAAGTTGATAACGGACTAGCCTTATTTTAACTTGCTATTTCTAGCTCTAAAAAAATTTCTTTTAGCTCTAAAACACCCGACTGTAAGGTGGTCCCGGCTGAT (50aa)
GGCTGGCTCAGGAGGGCGGAGGTCTCAGT-----GGGCGTCTGTAAGGTGGTCCCGGCT (43aa)
突变创建时间 2013-04-15
填表时间 2016/10/17
修改时间
PI/负责人 (工作单位) 孙永华(中国科学院水生生物研究所)
Genotype Information: