利用mRNA差异显示技术分离甜菜M14品系特异表达基因的cDNA片段

二倍体栽培甜菜与白花甜菜杂交、进一步回交而获得的单体附加系M₁₄，其染色体组成中除了含有18条栽培甜菜染色体外，还附加有一条野生白花甜菜第9号染色体，该附加染色体通过母本的传递率为96.5%；单体附加系传递率如此高的原因是因为M₁₄中有无融合生殖基因的存在。本实验采用mRNA差异展示技术对甜菜无融合生殖品系M₁₄和正常有性生殖的二倍体栽培甜菜A_2Y花蕾减数分裂时期的基因表达进行了差异分析。采用GT₁₅A，GT₁₅G，GT₁₅C 3种锚定引物，共筛选了20个随机引物，通过RT-PCR检测，获得了6个阳性差异表达的cDNA片段，应用NCBI的BLASTx软件对测序结果进行同源序列、相似序列检索,为进一步克隆无融合生殖基因提供侯选cDNA片段。

Extraction cDNA Fragments Specially Expressed in Lines M14 in Sugar Beet by mRNA Differential Display

A monosomic addition line in sugar beet, which was designated as M₁₄ was obtained by the wide cross between cultivated sugar beet (Beta vulgaris L.) and Beta corolliflora Zoss. M₁₄ contained the 18 normal chromosomes of sugar beet plus the B. corolliflora chromosome 9. This alien chromosome in M₁₄ had an average transmission frequency of 96.5% through eggs, the reason it had so high transmission frequency was that it had apomixis genes. By mRNA differential display method, the study compared gene expression at the stage of meiosis in florescence between M₁₄ and cultivated sugar beet (Beta vulgaris L.) A_2Y which can carry out normal sexual reproduction. The anchored primer were GT₁₅A, GT₁₅G, GT₁₅C,and twenty arbitrary primers were used. By RT-PCR, six positive specific cDNA fragments were obtained. Analysis was performed on NCBI, and comparison the similarity with published sequences was carried out using the BLASTx program. These cDNA fragments were the candidate fragments to clone the apomixis genes.