Figure 3.

RT-PCR Verification of Expression for Genes Cen3.t01107 and Cen3.t01160 in Cen3.

Gene Cen3.t01107 covers 3607 bp and has three splicing isoforms (Cen3.t01107.1 to Cen3.t01107.3) as predicted from existing transcript sequences. Two forward and two reverse primers were designed to differentiate these three isoforms; primer 113F was from an exon junction, but 113F/113R and 113F/114R still yielded very weak amplifications on genomic DNA for Cen3.t01107.1 and Cen3.t01107.3. Primer pair 113F/113R amplified the expected 383-bp fragment from all seven cDNA populations that corresponds to gene model Cen3.t01107.1, plus another two weak bands of 741 and ∼320 bp, respectively (arrows), while the 741-bp band represents the amplification product for gene model Cen3.t01107.3; the smaller band of 320 bp that is detected in five cDNA populations (not in cDNA samples from roots and buds) most likely represents a new isoform not yet identified. Gene models Cen3.t01107.2 and Cen3.t01107.3 are both transcribed in all seven tissues/treatments, with the sizes of the amplified products matching the annotation. Gene Cen3.t01160 has 11 exons totaling 7789 bp, and primers designed from exons 2 and 6 yielded amplification products in all seven cDNA samples. M, molecular marker; G, genomic DNA; L, cDNA from leaves; S, cDNA from shoots; R, cDNA from roots; P, cDNA from panicles; E, cDNA from etiolated leaves/shoots; B, cDNA from buds; C, cDNA from calli; N1 (L+S), N2 (R+P), and N3 (E+B+C), negative controls without adding reverse transcriptase (two or three negative controls for each primer pair were loaded to the same lane).