| ZKO number |
ZKO257 |
| Recent progress of this line |
F2 individuals have been indentified
|
|
Gene name |
TMEM66
|
|
Ensembl ID |
ENSDARG00000070452
|
|
Zebrafish strain |
AB
|
|
Cas9 target site 1 sequence |
antisense strand
GGGAACAGGGCTAGAGCGGCggg
|
|
Cas9 target site 2 sequence |
|
|
Position in the gene |
exon 2
|
|
Restriction enzyme (RE) |
n/a
|
|
PCR primer (forward) |
ATGCAAACCAATCCAACTCCTTTAT
|
|
PCR primer (reverse) |
AGATGCTGACTGTGGCTACTTCCTA
|
|
Length of expected PCR product (bp)
|
503
|
|
Length of expected RE digestion product (bp) |
|
|
Plasmids used for making Cas9 mRNA |
pXT7-hCAS9
|
| Plasmids used for making gRNA |
pT7-gRNA-ZKO257
|
| Primers used for making gRNA |
Pr-ZKO 257
|
|
Dosage of injected Cas9 mRNA (ng/ul)
|
200
|
|
Dosage of injected gRNA (ng/ul)
|
25
|
|
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/ZKO257.pdf
|
|
Mutation efficiency in F0 embryos (%)
|
90
|
Picture from F0 embryos
(gel picture or sequencing picture) |
|
|
Mutation type
|
Indel
|
|
Germline transmission efficiency in adult F0
|
4/7
|
|
Wild-type (WT) sequence
|
GGGAACAGGGCTAGAGCGGCgggCCGTGGT
|
Germline mutations in selected F0
(By screening F1 embryos) |
F0-1 |
|
|
F0-2
|
|
Mosaicism in selected F0
(By screening F1 adults) |
F0-1
|
A----------------------------CGGGCCGTGGT(-28bp)
|
|
F0-2
|
GGGAACAGGGCTAGAGCGGtcacGGCCGTGGT(-2bp,+4bp)
|
|
Allele name and predicted product
|
257a
257b |
A----------------------------CGGGCCGTGGT Truncated protein(178aa)
|
| GGGAACAGGGCTAGAGCGGtcacGGCCGTGGT Truncated protein(188aa) |
|
突变创建时间 |
2014-10-14
|
|
填表时间 |
2013/12/28
|
|
修改时间 |
2016/8/11
|
|
PI/负责人 (工作单位) |
|