Halfhill Matthew D, Good Laura L, Basu Chhandak, Burris Jason, Main Christopher L, Mueller Thomas C, Stewart C Neal
Department of Plant Sciences, University of Tennessee, Knoxville, TN 37996, USA.
Plant Cell Rep. 2007 Mar;26(3):303-11. doi: 10.1007/s00299-006-0219-1. Epub 2006 Oct 6.
The goal of this research was to generate a breeding population of horseweed segregating for glyphosate resistance. In order to generate a marker to select between hybrids of glyphosate resistant (GR) and glyphosate susceptible (GS) horseweed, a GR horseweed accession from western Tennessee was transformed with a green fluorescent protein (GFP) transgene. The GFP marker allowed for the simple and accurate determination of GR hybrid plants by visual observation. GR plants were shown to be transgenic via the green fluorescence under UV light, and resistant to glyphosate when sprayed with the field-use-rate 0.84 kg acid equivalent ha(-1) of glyphosate (i.e. Roundup) herbicide. An in vitro screen for glyphosate resistance in seedlings was developed, and a 5 microM glyphosate concentration was found to reduce dry weight in GS seedlings but not in GR seedlings. The GR plants containing GFP were then hand-crossed with GS plants from eastern Tennessee under greenhouse conditions, with GS plants acting as the pollen acceptor. Resulting seed was collected and germinated for GFP fluorescence screening. Seedlings that exhibited the transgenic GFP phenotype were selected as F(1) hybrids between GR and GS horseweed. Thirty GSxGR hybrids were produced on the basis of a green-fluorescent GFP phenotype of GR plants. GSxGFP/GR F(1) hybrids produced F(2) seeds, and F(2) plants were shown to segregate for GFP fluorescence and glyphosate resistance independently. Both traits segregated at a Mendelian 3:1 ratio, indicating a single gene is responsible for each phenotype.
本研究的目标是培育出对草甘膦抗性进行分离的马唐育种群体。为了生成一个用于在抗草甘膦（GR）和草甘膦敏感（GS）马唐的杂交种之间进行选择的标记，来自田纳西州西部的一个GR马唐种质用绿色荧光蛋白（GFP）转基因进行了转化。GFP标记使得通过视觉观察就能简单而准确地鉴定GR杂交植株。在紫外光下，GR植株通过绿色荧光显示为转基因植株，并且当用田间使用剂量0.84千克酸当量/公顷的草甘膦（即农达）除草剂进行喷洒时对草甘膦具有抗性。开发了一种用于幼苗草甘膦抗性的体外筛选方法，发现5微摩尔草甘膦浓度会降低GS幼苗的干重，但不会降低GR幼苗的干重。然后在温室条件下，将含有GFP的GR植株与来自田纳西州东部的GS植株进行人工杂交，GS植株作为花粉受体。收集产生的种子并进行萌发以进行GFP荧光筛选。表现出转基因GFP表型的幼苗被选为GR和GS马唐之间的F1杂交种。基于GR植株的绿色荧光GFP表型，产生了30个GS×GR杂交种。GS×GFP/GR F1杂交种产生了F2种子，并且F2植株显示出GFP荧光和草甘膦抗性是独立分离的。这两个性状均以孟德尔3:1的比例分离，表明每个表型由单个基因控制。
