中文摘要：甘蔗是在其10 Gb基因组中具有高多倍体和非整倍体的重要栽培作物。如果没有已知的拷贝数参考基因，便很难运用PCR技术精确估计甘蔗中任何基因的拷贝数。最近，有学者已研发出了可测量给定样本中目标DNA绝对量的液滴式数字PCR技术。本文利用液滴式数字PCR技术比较了澳大利亚三个转基因甘蔗的肌动蛋白解聚因子（ADF）、腺嘌呤磷酸核糖转移酶（APRT）和肌动蛋白（ACT）等内源基因的拷贝数。研究结果表明，液滴式数字PCR技术不仅可用于高通能量基因分析，而且是一种甘蔗基因拷贝数检测的快速、准确、可靠方法。
外文摘要：Sugarcane is an important cultivated crop with both high polyploidy and aneuploidy in its 10 Gb genome. Without a known copy number reference gene, it is difficult to accurately estimate the copy number of any gene of interest by PCR-based methods in sugarcane. Recently, a new technology, known as droplet digital PCR (ddPCR) has been developed which can measure the absolute amount of the target DNA in a given sample. In this study, we deduced the true copy number of three endogenous genes, actin depolymerizing factor (ADF), adenine phosphoribosyltransferase (APRT) and actin (ACT) in three Australian sugarcane varieties, using ddPCR by comparing the absolute amounts of the above genes with a transgene of known copy number. A single copy of the ACT allele was detected in Q208 (A) , two copies in Q240 (A) , but was absent in Q117. Copy number variation was also observed for both APRT and ADF, and ranged from 9 to 11 in the three tested varieties. Using this newly developed ddPCR method, transgene copy number was successfully determined in 19 transgenic Q208 (A) and Q240 (A) events using ACT as the reference endogenous gene. Our study demonstrates that ddPCR can be used for high-throughput genetic analysis and is a quick, accurate and reliable alternative method for gene copy number determination in sugarcane. This discovered ACT allele would be a suitable endogenous reference gene for future gene copy number variation and dosage studies of functional genes in Q208 (A) and Q240 (A) 
外文关键词：Droplet digital PCR (ddPCR); Sugarcane; Copy number; Brown rust resistance locus (Bru1)
作者：Sun, Yue; Joyce, Priya Aiyar
作者单位：Sugar Res Australia
期刊名称：PLANT CELL REPORTS
期刊影响因子：2.869
出版年份：2017
出版刊次：11
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