Assessing zygosity in progeny of transgenic plants: current methods and perspectives
Keywords:Digital PCR, homozygous line, LAMP assay, real-time PCR, transgenic plants
Homozygosity is highly desirable in transgenic plants research to ensure the stable integration and inheritance of transgene(s). Simple, reliable and high-throughput techniques to detect the zygosity of transgenic events in plants are invaluable tools for biotechnology and plant breeding companies. Currently, a number of basic techniques are being used to determine the zygosity of transgenic plants in T1 generation. For successful application of any technique, precision and simplicity of approach combined with the power of resolution are important parameters. On the basis of simplicity, resolution and cost involved, the available techniques have been classified into three major classes which are conventional methods, current methods and next generation methods. Conventional methods include antibiotic marker-based selection and the highly labour intensive Southern blot analysis. In contrast, methods such as real time PCR, TAIL PCR and competitive PCR are not only cost effective but rapid as well. Moreover, methods such as NGS, Digital PCR and Loop-mediated isothermal amplification also provide cost effective, fast and not so labour-intensive provide substitute of current methods. In this review we have attempted to compare and contrast all the available efficient methods to distinguish the homozygous plant in progeny of transgenics. This review also provides plenty of information regarding the various techniques available for determining zygosity in plants so as to permit the scientists to make informed choices of techniques that best suit their analyses. More importantly, detection and subsequent selection of homozygous individuals is central for facilitating the movement of transgenic plant from the laboratory to the field.
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