利用多重單一核苷酸基因定型技術鑑別國、內外食米品種
Authentication of Domestic Taiwan and Foreign Rice Varieties Based on Fingerprinting Analysis of Multiplex SNP Genotyping Technologies
Journal
作物、環境與生物資訊
Journal Volume
15
Journal Issue
3
Pages
184-197
Date Issued
2018
Author(s)
Abstract
為保持稻種純度，追溯、偵測市售混米，提昇國產稻米之競爭力及區隔性，稻米品種檢測技術之開發及推廣應用特別重要。除了以外觀、生理或生化特性進行稻種鑑定外，現今利用不同DNA分子標誌產生DNA指紋圖譜進行稻米品種鑑定已成為常用之分析技術。本研究室利用過去建立之國內外稻米品種單一核苷酸多型性(single nucleotide polymorphisms, SNPs)資料庫，篩選解析度最大之核心SNPs組，發展多重單一核苷酸基因定型（multiplex SNP genotyping）技術。此技術可以一般實驗室之分生配備，進行快速分析來鑑別台灣常見之市售國產米及國外進口食米品種。首先就不同品種間之SNP位點，以四引子擴增受阻突變體系聚合酶鏈反應(tetra-primer amplification refractory mutation system-polymerase chain reaction，tetra-primer ARMS-PCR)技術，分別設計適用於區分不同稉稻之13-plex與不同秈稻之10-plex引子對，再進行多重SNP基因定型反應，最後以一般或毛細管電泳比對DNA指紋圖譜確認品種。利用隨機取樣抽取單粒米DNA進行分析，可有效確認市售食米品種及混米率。此技術己完成專利申請並技術轉移中華穀類食品工業技術研究所。並已應用於擴增國、內外水稻品種DNA資料庫，市售稻米品種確認，田間異品種及不同生產階段稻米混雜率之檢出。此技術將有助於國市場食米品質管制機制之建立，維護消費者之權益及信心，進而有益台灣稻米之發展。
In order to maintain genetic purity of rice seeds, detect and trace admixtures of commercial rice, increase the competitiveness and marketing segmentation of domestic Taiwan rice, the development and promotion of rice variety identification technology is particularly important. For the rice variety identification, except by outside appearance, physiological or biochemical characteristics, currently the DNA fingerprinting technique based on different DNA molecular markers has become a commonly used platform. In this study, we took advantage of a core SNP marker set with highest resolution from previously rice SNP database to establish different genotyping analysis platforms for distinguishing domestic and foreign rice varieties. This technology can be simply adopted by general laboratory with molecular biology equipment for rapid analysis to identify common domestically produced Taiwan rice and imported rice varieties. Firstly, the tetra-primer amplification refractory mutation system-polymerase chain reaction (tetra-primer ARMS-PCR) technique was used to design a 13-plex and 10-plex allelic-specific SNP primer pairs based on specific SNP loci among various rice cultivars. Then, multiple SNP genotyping were performed to distinguish japonica and indica rice varieties, respectively; and the PCR amplicons were analyzed by capillary or regular gel electrophoresis. Finally, the identity of variety was confirmed by comparing the DNA fingerprinting profiling with available rice SNP database. Our results showed that the use of random sampling to extract single-grain DNA for multiple SNP genotyping analysis can effectively confirm the varieties of rice and the mixed rice rate. This technology has completed patent application and technology transfer to China Grain Products Research and Development Institute. In addition, this technology has been applied to the amplification of DNA databases for domestic and foreign rice varieties, the confirmation of commercial rice, the detection of off-type rice in field, and the monitoring of mixed rice rates at different production stages. This technology will be helpful to set up the quality control of commercial market system, maintain consumer rights and confidence, and benefit the development of rice industry in Taiwan.
Subjects
四引子擴增受阻突變體系聚合酶鏈反應 ； 單一核苷酸多型性 ； 多重基因定型 ； DNA指紋圖譜 ； 食米品種鑑定 ；
Tetra-primer ARMS-PCR ； single nucleotide polymorphism (SNP) ； multiplex SNP genotyping ； DNA fingerprinting profile ； rice varieties identification
Publisher
臺灣農藝學會 & 中華農業氣象學會
Type
journal article