Xenbase Image ID: 156741

Fig. 4. Loss-of-function study of xBest-2. (A) Genomic structures including exon 1 (1) and exon 2 (2) of xBest-2a and xBest-2b. The translation initiation codon (ATG) and primer regions are indicated (Primer). The splice-site-targeted antisense morpholino oligonucleotides, xBest-2a MO and xBest-2b MO, were designed against the sequence at the boundary of exon 1/intron 1 of each gene. xBest-2a and xBest-2b intron 1 have different lengths. The pre-mRNAs of xBest-2a and xBest-2b generated by splice blocking have a termination codon immediately following exon 1. The intron 1 sequences are in lower case. Amino acid sequences are also indicated. (B) RT-PCR analysis of xBest-2 mRNA in MO-injected embryos. Embryos were injected with 20 ng of xBest-2a MO plus 20 ng of xBest-2b MO (B2), or 40 ng of standard control MO (SC), in the marginal region of both blastomeres at the 2-cell stage, and cultured for RT-PCR until various stages as indicated above each lane. The primer set was designed to span the first intron to detect splicing of the endogenous pre-mRNA for both xBest-2a and xBest-2b (corresponding to Primer in (A)). In xBest-2 MO-injected embryos, the mature mRNA (m) was decreased and nonspliced pre-mRNA (p1 and p2) was detected from stage 10.5. The mature xBest-2 mRNA was eliminated at stage 30 in xBest-2 MO-injected embryos. Splicing of xBest-2 pre-mRNA was not affected by control MO injection. (C) A control uninjected embryo at stage 35 (uninjected). (D) An embryo injected with 20 ng of xBest-2a MO plus 20 ng of xBest-2b MO (xBest-2 MOs). Disruption of xBest-2 did not affect the patterning of embryos at stage 35. Image published in: Onuma Y et al. (2009) Copyright © 2009. This image is reproduced 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. Larger Image Printer Friendly View

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