High capacity thermal cycler for molecular genotyping

拟南芥植物hth基因的隐性突变等位基因纯合可以继承祖先的DNA序列信息，不存在于其父母的染色体基因组。这种现象是全基因组的，并且发生在基因组的编码区和非编码区。这些发现是全新的，提出的机制也是前所未有的。 这一发现在很大程度上是由于分子DNA技术的进步。随着PCR技术的出现、拟南芥基因组测序的完成和寡核苷酸合成方法的进步，现在快速可靠地对大量个体进行基因分型变得微不足道。 因为这些分子标记是共显性的，所以也可以同时对两条染色体进行采样。 因此，鉴定携带两个相同等位基因或两个序列仅相差一个核苷酸的不同等位基因的个体是非常简单的。在本申请中，要求资金购买高容量模块化热循环仪，用于对突变型热头植物及其后代的群体进行基因分型。 我建议使用PCR技术和分子标记来检测基因组编码区和非编码区DNA序列的变化。我们将使用这些简单的测定来阐明这些突变体中发现的新型遗传不稳定性的显著特征。 如果这一过程在植物界以外的生物体中发现，这项研究可能在未来具有重要的医学意义。

Arabidopsis thaliana plants homozygous for recessive mutant alleles of the hothead (hth) gene can inherit ancestral DNA sequence information that was not present in the chromosomal genome of their parents. This phenomenon is genome-wide and occurs in both coding and non-coding regions of the genome. These findings are completely novel and the mechanism proposed, unprecedented.  This discovery was made possible in large part by advances in molecular DNA technology. With the advent of PCR, the completion of the Arabidopsis genome sequence and advances in oligonucleotide synthesis procedures, it is now trivial to genotype large numbers of individuals rapidly and reliably.  Because these molecular markers are co-dominant it is also possible to sample both chromosomes simultaneous.  Hence, identifying individuals that carry two alleles that are identical or two distinct alleles that differ in sequence by only one nucleotide is very straightforward. In this application funds are requested to purchase a high capacity modular thermal cycler for genotyping populations of mutant hothead plants and their descendents.  I propose to use PCR technology and molecular markers to assay for changes in DNA sequences  in coding as well as  in non-coding regions of the genome. We will use these simple assays to elucidate the salient features of the novel type of genetic instability uncovered in these mutants.  If this process is found in organisms outside of the plant kingdom, it is possible that this research will have significant future medical relevance.