蛋白表達穩定性的影響因素

外源蛋白在不同遺傳背景、不同生長環境、不同發育時期和不同的組織類型下表達的穩定性是轉基因作物性狀穩定性的根本。為了評估基因型、生長地、發育階段等因素對蛋白表達的影響，杜邦先鋒公司的研究人員以玉米中的3個內源看家蛋白作為研究對象，系統分析了各種因素對其表達穩定性的影響。

看家蛋白參與了基本的細胞學過程，其表達相對來講比較穩定，因此常作為內參對實驗系統進行平衡化處理。研究不同環境下看家蛋白的表達變化能夠間接反映出轉基因作物中外源蛋白受基因型、生長地、發育階段等因素的影響程度。研究人員在印度多個試驗地點種植了一系列的雜交玉米材料，並連續兩年通過ELISA實驗測定了3個發育時期的葉片中3個看家蛋白的含量。這3個看家蛋白選自細胞骨架結構相關的Actin蛋白、氨基酸鏈延伸相關的EF1-alpha（Elongation Factor 1-alpha）蛋白和糖酵解途徑中的GAPDH（glyceraldehyde 3-phosphate dehydrogenase）蛋白。結果表明，玉米內源蛋白的表達受到基因型的影響較小，而受到生長環境的影響較大。

該研究對於系統評估轉基因作物的環境適應性和穩定性具有極大的借鑒意義。

Transgenic Research, 17 May 2018

Expression of endogenous proteins in maize hybrids in a multi-location field trial in India

Authors

Linga R. Gutha, Divakar Purushottam, Aruna Veeramachaneni……Jennifer Anderson*, Bonnie Hong, Jean Schmidt, Shveta Bagga

*: DuPont Pioneer, USA

Abstract

Genetically modified (GM) crops undergo large scale multi-location field trials to characterize agronomics, composition, and the concentration of newly expressed protein(s) [herein referred to as transgenic protein(s)]. The concentration of transgenic proteins in different plant tissues and across the developmental stages of the plant is considered in the safety assessment of GM crops. Reference or housekeeping proteins are expected to maintain a relatively stable expression pattern in healthy plants given their role in cellular functions. Understanding the effects of genotype, growth stage and location on the concentration of endogenous housekeeping proteins may provide insight into the contribution these factors could have on transgenic protein concentrations in GM crops. The concentrations of three endogenous proteins (actin, elongation factor 1-alpha, and glyceraldehyde 3-phosphate dehydrogenase) were measured in several different maizehybrids grown across multiple field locations over 2 years. Leaf samples were collected from healthy plants at three developmental stages across the growing seasons, and protein concentrations were quantified by indirect enzyme-linked immunosorbent assay (ELISA) for each protein. In general, the concentrations of these three endogenous proteins were relatively consistent across hybrid backgrounds, when compared within onegrowth stage and location (2–26%CV), whereas the concentrations of proteins in the same hybrid and growth stage across different locations were more variable (12–64%CV). In general, the protein concentrations in 2013 and 2014 show similar trends in variability. Some degree of variability in protein concentrations should be expected for both transgenic and endogenous plant-expressed proteins. In the case of GM crops, the potential variation in protein concentrations due to location effects is captured in the current model of multi-location field testing.

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