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Improvement of the dot-blot-SNP technique for efficient and cost-effective genotyping

机译：改进点印迹SNP技术以进行有效和具有成本效益的基因分型

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Although the dot-blot-SNP technique is a laborsaving, cost-effective method for SNP genotyping of a large number of plants, the synthesis of 5o-digoxigenin (DIG)-labeled oligonucleotides for use as probes is still costly. We developed two probe-labeling methods for this technique, one being digoxigenin labeling of oligonucleotides by PCR (PCR-DIG labeling) and the other being hybridization using a bridge probe and a 5o-DIG-labeled oligonucleotide (bridge hybridization). Bridge hybridization detected allele-specific signals under hybridization conditions similar to those for the 5o-DIG-labeled oligonucleotides and biotin-labeled oligonucleotides, while signals were detected only under a lower stringency condition by PCR-DIG labeling. As a method for genotyping using many markers at one time, two methods, i.e., PCR using mixed primer pairs and hybridization using mixed probes, were examined with successful results. Eighty-five SNP markers designed for genotyping of rice cultivars detected allele-specific signals, the genotyping results corresponding to the previously reported ones.
机译：尽管斑点印迹SNP技术是一种用于大量植物SNP基因分型的省力，经济高效的方法，但用作探针的5o-地高辛配基（DIG）标记的寡核苷酸的合成仍然昂贵。我们开发了两种用于该技术的探针标记方法，一种是通过PCR对寡核苷酸进行洋地黄毒苷标记（PCR-DIG标记），另一种是使用桥探针和5o-DIG标记的寡核苷酸进行杂交（桥杂交）。桥接杂交在类似于5o-DIG标记的寡核苷酸和生物素标记的寡核苷酸的杂交条件下检测到等位基因特异性信号，而信号仅通过PCR-DIG标记在较低严格条件下检测到。作为一次使用许多标记进行基因分型的方法，研究了两种方法，即使用混合引物对的PCR和使用混合探针的杂交，均获得了成功的结果。设计用于水稻品种基因分型的八十五个SNP标记物检测到等位基因特异性信号，其基因分型结果与先前报道的信号相对应。

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