ICAR-Indian Agricultural Research Institute, Pusa, New Delhi, 110 012, India.
CSIR-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, 176 061, India.
Biol Futur. 2020 Jun;71(1-2):165-173. doi: 10.1007/s42977-020-00014-8. Epub 2020 Jun 11.
Brassica oleracea cv. Pride of India is one of the most promising vegetable cultivars commercially grown as cash crop in Himachal Pradesh, India. However, its overall production is severely hampered by diamondback moth (Plutella xylostella), a notorious pest. To avoid yield losses caused by this pest, cryIAa gene was introduced into cabbage cv. Pride of India using Agrobacterium-mediated transformation method. In an attempt to maximize the transformation frequency, critical determinant factors such as explant type, pre-incubation and co-cultivation period, and acetosyringone effect were successfully optimized. The highest transformation frequency (4.67% and 14.50%) in cotyledon and hypocotyl explant was achieved with a pre-incubation period of 72 h and co-cultivation period of 48 h. Furthermore, transformation frequency was enhanced in cotyledon (18.66%) and hypocotyl (32.00%) explants, when selective regeneration medium was fortified with 100 µM acetosyringone, respectively. The transgene (cryIAa) integration and copy number were confirmed using PCR and Southern blotting. Reverse transcriptase PCR and quantitative real-time PCR analyses were performed that proved transcriptional expression of cryIAa gene in PCR-positive transgenic events. Transgenic cabbage-fed diamondback moth larvae showed significantly higher mortality, thereby proving transgene effectiveness against insect pest control.
印度甘蓝型白菜 cv. Pride of India 是印度喜马偕尔邦商业种植的最有前途的蔬菜品种之一，是一种经济作物。然而，其总产量严重受到小菜蛾（Plutella xylostella）的阻碍，小菜蛾是一种臭名昭著的害虫。为了避免这种害虫造成的产量损失，将 cryIAa 基因通过农杆菌介导的转化方法导入白菜 cv. Pride of India。为了最大限度地提高转化频率，成功优化了外植体类型、预培养和共培养时间以及乙酰丁香酮效应等关键决定因素。在预培养 72 h 和共培养 48 h 的条件下，子叶和下胚轴外植体的最高转化频率（4.67%和 14.50%）。此外，在选择性再生培养基中添加 100 µM 乙酰丁香酮，分别提高了子叶（18.66%）和下胚轴（32.00%）外植体的转化频率。利用 PCR 和 Southern blot 证实了转基因（cryIAa）的整合和拷贝数。反转录 PCR 和定量实时 PCR 分析证明了 cryIAa 基因在 PCR 阳性转基因事件中的转录表达。喂食转基因白菜的小菜蛾幼虫死亡率明显更高，从而证明了转基因在防治虫害方面的有效性。
