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XB-IMG-84552

Xenbase Image ID: 84552


Fig. 5. gRNA/Cas9-mediated gene targeting is suitable for G0 phenotyping. (A-L) Whole-mount in situ hybridization analysis of expression of the pancreas-specific marker pdip in X. tropicalis normal control tadpoles (stage 40), tadpoles injected with Cas9 mRNA (300 pg/embryo) and gRNAs (50 pg/embryo), and tadpoles injected with either ptf1a/p48GR mRNA (20 pg/embryo) alone or in combination with Cas9 and ptf1a/p48 gRNA. Dexamethasone (working concentration of 10 μM) was added at stage 14 to activate Ptf1a/p48. (A,E) Complete inhibition of pdip expression upon targeting ptf1a/p48. (B,F) Partial inhibition of pdip expression upon targeting ptf1a/p48. (C,G) Overexpression of ptf1a/p48 expands pdip expression in the territory of the stomach and duodenum. (D,H) The inhibition of pdip expression upon gRNA/Cas9-mediated targeting of ptf1a/p48 was completely rescued by co-injection of ptf1a/p48GR mRNA. (I,K) Uninjected control embryos. (J,L) As a negative control, pdip expression was unaffected upon targeting elastase. All the images are lateral views with the head to the left. E-H,K,L show further examples of the types represented in A-D,I,J, respectively. The number of embryos showing the illustrated phenotype is given in the representative image. (M,N) Dissection of 1-week-old froglets revealed severe pancreatic hypoplasia in ptf1a/p48 gRNA/Cas9-injected G0 froglets, with stomach and duodenum unaffected. The pancreas is outlined (dashed line). du, duodenum; st, stomach. (O-V) Albinism phenotype caused by tyrosinase gRNA/Cas9. (O,S) Uninjected control tadpoles. (P,T) Almost full albinism. (Q,U) Tadpoles showing severe perturbation of pigmentation. (R,V) Partial albinism. S-V show further examples of the types represented in O-R, respectively. The number of embryos showing the illustrated phenotype is given in the representative image. (W-Z) Dorsal view of adult frogs. (W) Wild type. (X) Almost full albinism caused by tyrosinase gRNA/Cas9. (Y,Z) Partial albinism. The numbers of frogs showing the illustrated phenotypes are listed. Scale bars: 400 μm in A-L; 2 mm in M,N; 1 mm on O-V; 1 cm in W-Z.

Image published in: Guo X et al. (2014)

Copyright © 2014. Image reproduced on Xenbase with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License.

GeneCloneSynonymsSpeciesStage(s)Tissue
pdia2 pdi, XPDIp, pdipX.tropicalisThroughout NF stage 40dorsal pancreatic bud
ventral pancreatic bud
pancreas

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