Department of Plant Sciences, University of Tennessee, Knoxville, TN 37996, USA.
Biotechnol J. 2011 Jan;6(1):118-23. doi: 10.1002/biot.201000258.
Assaying for transgenic pollen, a major vector of transgene flow, provides valuable information and essential data for the study of gene flow and assessing the effectiveness of transgene containment. Most studies have employed microscopic screening methods or progeny analyses to estimate the frequency of transgenic pollen. However, these methods are time-consuming and laborious when large numbers of pollen grains must be analyzed to look for rare transgenic pollen grains. Thus, there is an urgent need for the development of a simple, rapid, and high throughput analysis method for transgenic pollen analysis. In this study, our objective was to determine the accuracy of using flow cytometry technology for transgenic pollen quantification in practical application where transgenic pollen is not frequent. A suspension of non-transgenic tobacco pollen was spiked with a known amount of verified transgenic tobacco pollen synthesizing low or high amounts of green fluorescent protein (GFP). The flow cytometric method detected approximately 75% and 100% of pollen grains synthesizing low and high amounts of GFP, respectively. The method is rapid, as it is able to count 5000 pollen grains per minute-long run. Our data indicate that this flow cytometric method is useful to study gene flow and assessment of transgene containment.
对转基因花粉进行分析，作为转基因流的主要载体，为基因流研究和评估转基因控制的有效性提供了有价值的信息和必要的数据。大多数研究采用显微镜筛选方法或后代分析来估计转基因花粉的频率。然而，当需要分析大量花粉粒以寻找罕见的转基因花粉粒时，这些方法既费时又费力。因此，迫切需要开发一种简单、快速和高通量的转基因花粉分析方法。在这项研究中，我们的目的是确定使用流式细胞术技术对转基因花粉进行定量分析的准确性，特别是在转基因花粉不频繁的实际应用中。将一定量经验证的合成低或高量绿色荧光蛋白（GFP）的转基因烟草花粉悬浮液混入非转基因烟草花粉中。流式细胞术方法分别检测到合成低和高量 GFP 的花粉粒约为 75%和 100%。该方法快速，因为它能够在每分钟运行中计数 5000 个花粉粒。我们的数据表明，这种流式细胞术方法可用于研究基因流和评估转基因控制。
