| ZKO number |
ZKO33 |
| Recent progress of this line |
F2 individuals have been indentified
|
|
Gene name |
sc:d0144
|
|
Ensembl ID |
ENSDARG00000040543
|
|
Zebrafish strain |
AB
|
|
Cas9 target site 1 sequence |
antisense strand
GGCGGCGGGGTGAAGGCGCAggg
|
|
Cas9 target site 2 sequence |
|
|
Position in the gene |
Exon 2
|
|
Restriction enzyme (RE) |
n/a
|
|
PCR primer (forward) |
CCCGCAGAAGCAGTATAAGGTA
|
|
PCR primer (reverse) |
AGTCAGATCAAACGTCAGACCC
|
|
Length of expected PCR product (bp)
|
415
|
|
Length of expected RE digestion product (bp) |
49+366
|
|
Plasmids used for making Cas9 mRNA |
pSP6-2sNLS-spCas9
|
| Plasmids used for making gRNA |
pT7-gRNA-ZKOpT7-gRNA-ZKO33
|
| Primers used for making gRNA |
Pr-ZKO 33
|
|
Dosage of injected Cas9 mRNA (ng/ul)
|
300
|
|
Dosage of injected gRNA (ng/ul)
|
75
|
|
Survival rate of F0 embryo @ 1dpf (%)
|
92.5
|
|
Method to estimate mutation efficiency |
T7EI detection
|
|
Stage of the embryos collected for mutation detection |
http://www.zfish.cn/upload/file/ZKOPDF/ZKO33.pdf
|
|
Mutation efficiency in F0 embryos (%)
|
15
|
Picture from F0 embryos
(gel picture or sequencing picture) |
|
|
Mutation type
|
Indel
|
|
Germline transmission efficiency in adult F0
|
60% (m/n: Number of positive F0 / Total number of F0 screened)
|
|
Wild-type (WT) sequence
|
TGCCCTGCGCCTTCACCCCGCCG
|
Germline mutations in selected F0
(By screening F1 embryos) |
F0-1 |
TGCCCTxxxxxxTCAxxCCxCCG ( -10 bp)
|
|
F0-2
|
TGCxxxxxxCCTTCACCCCGCCG (-6 bp)
|
Mosaicism in selected F0
(By screening F1 adults) |
F0-1
|
25% (m/n: Number of F1 carrier / Total number of F1 screened)
CGCCGxxxGTCGGATCC (-3 bp)
|
|
F0-2
|
31.25% (m/n: Number of F1 carrier / Total number of F1 screened)
CCTxxxxxxTCAxxCCxCCGCCG ( -14bp,+5bp)
|
|
Allele name and predicted product
|
33a
33b |
CCTxxxxxxTCAxxCCxCCGCCG, delete three aa(92nd,93rd,94th) and C90F,A91T
|
| |
|
突变创建时间 |
2014-09-01
|
|
填表时间 |
2013/12/24
|
|
修改时间 |
2016/8/8
|
|
PI/负责人 (工作单位) |
|