Multiple Transgenic Strategies Positively Regulate Cold-Induced Sweetening in Low Temperature Stored Potato Tubers

IF 2.3 3区 农林科学 Q1 AGRONOMY
Nida Toufiq, Olawale Samuel Adeyinka, Anwar Khan, Shazia Shafique, Nusrat Jahan, Muhammad Umar Bhatti, Rida Khalid, Ayman Naeem, Qamar Abbas, Sobiya Shafique, Bushra Tabassum
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Abstract

Cold-induced sweetening (CIS) is a common phenomenon in many plants including potatoes that help in osmoregulation and cryoprotection. However, CIS is associated with quality deterioration in potato tubers due to accumulation of reducing sugars at low temperatures. We investigated two different strategies to modulate CIS in potato, overexpression of RING finger (SbRFP1) as anti-sweetening gene and by double-stranded RNA-mediated gene silencing of the vacuolar invertase gene. In silico analysis predicted that the ubiquitination activity of the RING finger protein was responsible for regulating the expression of invertase during cold-induced stress. Moreover, the in silico predicted binding stability of siRNA-mRNA duplex suggested efficient gene silencing of the invertase gene. We successfully generated four single and three dual transgenic potato lines that were positive for transgene insertion and integration as revealed in PCR and Southern blot. The amount of reducing sugars found in tubers obtained from single transgenics showed maximum decrease of 1.67 folds while tubers obtained from dual transgenic line depicted 4.86 folds reduced accumulation of reducing sugars compared to non-transgenic control when analyzed through HPLC analysis post 60-day storage at low temperature (4°C). Further, the invertase activity was 1.46 folds reduced in single transgenics while this reduction was 2.13 folds in dual transgenics. The downregulation of the invertase gene was up to 3.36 folds in dual transgenic potato lines, 2.26 folds in single transgenic compared to control, non-transgenic post 60-day cold storage at low temperature. Conclusively, the utilization of multiple strategies to regulate CIS in low-temperature stored potato tubers positively regulate CIS in transgenic potatoes and can be employed to generate CIS resistant potato varieties.

Abstract Image

多种转基因策略积极调控低温贮藏马铃薯块茎的冷诱导甜度
冷诱导增甜(CIS)是包括马铃薯在内的许多植物的常见现象,有助于渗透调节和低温保护。然而,由于还原糖在低温下积累,CIS 与马铃薯块茎的品质下降有关。我们研究了两种不同的策略来调节马铃薯的 CIS,一种是过量表达 RING 手指(SbRFP1)作为抗变甜基因,另一种是通过双链 RNA 介导的液泡转化酶基因沉默。硅学分析预测,RING 手指蛋白的泛素化活性在冷诱导胁迫过程中负责调节转化酶的表达。此外,硅学预测的 siRNA-mRNA 双链体的结合稳定性表明可以有效地沉默转化酶基因。我们成功培育出了四个单转基因马铃薯品系和三个双转基因马铃薯品系,PCR和Southern印迹显示这些品系的转基因插入和整合均呈阳性。在低温(4°C)贮藏 60 天后,通过 HPLC 分析,单转基因块茎中的还原糖含量比非转基因对照减少了 4.86 倍。此外,单一转基因品系的转化酶活性降低了 1.46 倍,而双转基因品系则降低了 2.13 倍。在低温冷藏 60 天后,与对照和非转基因相比,双转基因马铃薯品系的转化酶基因下调高达 3.36 倍,单转基因为 2.26 倍。总之,利用多种策略调节低温贮藏马铃薯块茎中的转化酶基因对转基因马铃薯的转化酶基因有积极的调节作用,可用于培育抗转化酶基因的马铃薯品种。
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来源期刊
Potato Research
Potato Research AGRONOMY-
CiteScore
5.50
自引率
6.90%
发文量
66
审稿时长
>12 weeks
期刊介绍: Potato Research, the journal of the European Association for Potato Research (EAPR), promotes the exchange of information on all aspects of this fast-evolving global industry. It offers the latest developments in innovative research to scientists active in potato research. The journal includes authoritative coverage of new scientific developments, publishing original research and review papers on such topics as: Molecular sciences; Breeding; Physiology; Pathology; Nematology; Virology; Agronomy; Engineering and Utilization.
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