High efficiency transformation of Brassica oleracea var. botrytis plants by Rhizobium rhizogenes

Abstract

Brassica oleracea var. botrytis, a very popular crop grown for its edible inflorescence, is bred only as a mutated annual cultivar and does not naturally occur in environment. Since cauliflower is still described as the most troublesome of all the B. oleracea vegetables regarding transformation processes, it is fully justified to focus on the improvement of tools for its genetic modifications. Here, we present a successful protocol for genetic transformation of cauliflower employing the process of agroinfection. The primary analysis of in vitro response of five cultivars allowed us to have chosen Pionier as the most promising cultivar; in consequence the Pionier was transformed via Rhizobium-mediated techniques in order to evaluate both, R. radiobacter (EHA 105, LBA 4404) and R. rhizogenes (ATCC 18534, A4) species. However, the latter system turned out to be more effective and, the A4 strain, in particular (72% transformation efficiency, 55% confirmed by GUS assay). That shows a promising technical advance especially when compared to the results of previous literature reports (e.g. 8.7% reported efficiency using R. rhizogenes). The transgenic cauliflower was obtained from hairy roots via organogenic callus induction. The potential transformants were analysed at the genomic and proteomic levels and their transgenic character was fully confirmed.Electronic supplementary materialThe online version of this article (10.1186/s13568-018-0656-6) contains supplementary material, which is available to authorized users.

Establishment of Agrobacterium-mediated genetic transformation and application of CRISPR/Cas9 gene-editing system to Chinese cabbage (Brassica rapa L. ssp. pekinensis)

“…Among 123 genotypes of Chinese cabbage, a large variation in regeneration frequency, ranging from 0-95%, was observed [14]. Hypocotyls are usually selected as explants for regeneration and transformation in Brassica crops [3,35,37]. However, in this study, hypocotyl explants expanded to form calli only.…”

“…Stable and high-efficiency transformations have been conducted in a variety of Brassica crops, particularly B. napus and B. oleracea [2,35,36]. This study identified various factors that influence Agrobacterium-mediated plant transformation in Chinese cabbage.…”

Breeding for Nutritional and Organoleptic Quality in Vegetable Crops: The Case of Tomato and Cauliflower

“…The pSAG12:ipt gene was evidenced to be responsible for delay in leaf senescence, altered synchronous curd initiation, smaller curd size, and higher susceptibility to fungal infection [135]. More recently, Kowalczyk et al (2018) developed a high efficiency cauliflower transformation method using Rhizobium (=Agrobacterium) rhizogenes in hypocotyl sections of 10-day-old de-rooted seedling [136]. Zhou and co-workers (2008) isolated the Or gene from orange cauliflower and introduced the transgene in potato revealing that the gene controls carotenoid accumulation by inducing the formation of chromoplasts [137].…”

Development of Transgenic Brassica Crops against Biotic Stresses Caused by Pathogens and Arthropod Pests

“…With regard to Agrobacterium tumefaciens, effective transformations in cotyledonary or hypocotyl explants of Chinese cabbage [15,16], or by floral dip in rapeseed [17] have been reported. In addition, there are perfectly developed protocols for this from seedling explants [18], even having developed an effective transient expression system [19], and also with Agrobacterium rhizogenes [20]. However, nowadays, methods are beginning to be developed with other vector bacteria, such as the rhizospheric bacterium of rapeseed Ensifer adhaerens, which is capable of genetically transforming its original host [21].…”