Cnudde Filip, Hedatale Veena, de Jong Hans, Pierson Elisabeth S, Rainey Daphne Y, Zabeau Marc, Weterings Koen, Gerats Tom, Peters Janny L
Institute for Wetland and Water Research, Department of Experimental Botany, Section Plant Genetics, Radboud University, Toernooiveld 1, 6525 ED, Nijmegen, The Netherlands.
Chromosome Res. 2006;14(8):919-32. doi: 10.1007/s10577-006-1099-5. Epub 2007 Jan 19.
We analyzed changes in gene expression during male meiosis in Petunia by combining the meiotic staging of pollen mother cells from a single anther with cDNA-AFLP transcript profiling of mRNA from the synchronously developing sister anthers. The transcript profiling experiments focused on the identification of genes with a modulated expression profile during meiosis, while premeiotic archesporial cells and postmeiotic microspores served as a reference. About 8000 transcript tags, estimated at 30% of the total transcriptome, were generated, of which around 6% exhibited a modulated gene expression pattern at meiosis. Cluster analysis revealed a transcriptional cascade that coincides with the initiation and progression through all stages of the two meiotic divisions. Fragments that exhibited high expression specifically during meiosis I were characterized further by sequencing; 90 out of the 293 sequenced fragments showed homology with known genes, belonging to a wide range of gene classes, including previously characterized meiotic genes. In-situ hybridization experiments were performed to determine the spatial expression pattern for five selected transcript tags. Its concurrence with cDNA-AFLP transcript profiles indicates that this is an excellent approach to study genes involved in specialized processes such as meiosis. Our data set provides the potential to unravel unique meiotic genes that are as yet elusive to reverse genetics approaches.
我们通过将单个花药中花粉母细胞的减数分裂分期与同步发育的姐妹花药中mRNA的cDNA-AFLP转录谱相结合，分析了矮牵牛雄配子减数分裂过程中的基因表达变化。转录谱实验着重于鉴定减数分裂过程中表达谱发生变化的基因，而减数分裂前的孢原细胞和减数分裂后的小孢子作为对照。共产生了约8000个转录标签，估计占转录组总量的30%，其中约6%在减数分裂时表现出基因表达模式的变化。聚类分析揭示了一个转录级联反应，它与减数分裂两个阶段的起始和进程相吻合。对在减数分裂I期间特异性高表达的片段进行了进一步测序分析；在293个测序片段中，有90个与已知基因具有同源性，这些基因属于广泛的基因类别，包括先前已鉴定的减数分裂基因。我们进行了原位杂交实验，以确定五个选定转录标签的空间表达模式。其与cDNA-AFLP转录谱的一致性表明，这是研究参与减数分裂等特殊过程的基因的一种优秀方法。我们的数据集为揭示反向遗传学方法尚未发现的独特减数分裂基因提供了可能。
