Figure 3

Alternative splice variants of the zebrafish NXT2. A: Solid black boxes represent exons, lines represent introns. Initiation codons and stop codons are indicated in capitalized letters. Solid blue box represents an exon of human NXT2 splice variant NXT2b, with GTG as initiation codon. The zebrafish NXT2 splice variant NXT2b is indicated below zebrafish NXT2 with GTG as initial codon. Primers NTf/NTr and NXTf/NXTrm (horizontal arrows) were used for PCR amplification to identify splicing variants of zNXT2. chr.: chromosome or linkage group. B: RNA expression profile of zebrafish NXT2 splice variants by RT-PCR. Equal amounts (2 μg) of total RNA, isolated from wild-type embryos (lanes 1, 4, 7), NXT2 mutant embryos (lanes 3, 6, 9) and their heterozygous siblings (lanes 2, 5, 8), were used for RT-PCR analyses. Lanes 1–3 refer to PCR products of endogenous eflα gene as positive control, lanes 4–6 represent products of primers NXTf/NXTrm, and lanes 7–9 indicate PCR products of primers NTf/NTr. RNA transcription level of zNXT2, represented by NTf/NTr in lane 9 (indicated by an asterisk), is remarkably reduced in mutant embryos, compared with that of their heterozygous and wild-type sibling embryos in lanes 8 and 7. No significant difference is detected among different genotypic embryos by NXTf/NXTrm in lanes 4, 5 and 6. -/-: zNXT2 homozygous mutant; +/-: heterozygous sibling;+/+: wild type sibling; ef1 alpha, NXT2a and NXT2b: transcripts detected by appropriate primers.