Efficient sweet pepper transformation mediated by the BABY BOOM transcription factor

Abstract

Pepper (Capsicum L.) is a nutritionally and economically important crop that is cultivated throughout the world as a vegetable, condiment, and food additive. Genetic transformation using Agrobacterium tumefaciens (agrobacterium) is a powerful biotechnology tool that could be used in pepper to develop community-based functional genomics resources and to introduce important agronomic traits. However, pepper is considered to be highly recalcitrant for agrobacterium-mediated transformation, and current transformation protocols are either inefficient, cumbersome or highly genotype dependent. The main bottleneck in pepper transformation is the inability to generate cells that are competent for both regeneration and transformation. Here, we report that ectopic expression of the Brassica napus BABY BOOM AP2/ERF transcription factor overcomes this bottleneck and can be used to efficiently regenerate transgenic plants from otherwise recalcitrant sweet pepper (C. annuum) varieties. Transient activation of BABY BOOM in the progeny plants induced prolific cell regeneration and was used to produce a large number of somatic embryos that could be converted readily to seedlings. The data highlight the utility of combining biotechnology and classical plant tissue culture approaches to develop an efficient transformation and regeneration system for a highly recalcitrant vegetable crop.

Fig. 1

Regeneration response of sweet pepper explants. Response of sweet pepper ‘Fiesta’ explants to co-cultivation with agrobacterium carrying either the 35S:GUS construct (a–c) or the 35S::BnBBM:GR construct (d–f). a Callus formation (arrow) and shoot-like structures (SLS; asterisk) after 3 weeks of culture; b leaf-like structures developing at the wounded edge of a cotyledon after 4 weeks of culture; c explant with callus (arrow) and SLS (asterisk) histochemically stained for GUS activity; d SLS, 4 weeks after treatment; e somatic embryo formation (arrows) on a newly emerged leaf, after 6 weeks of culture; f elongating SLS (asterisk) after 4 weeks on elongation medium

Fig. 2

Regeneration response of stable 35S::BBM:GR sweet pepper lines. a–d 24-day-old seedlings germinated on medium with the indicated supplements; e–h feather-cut cotyledons of 10-day-old seedlings incubated for 14 days on medium with the indicated supplements. Shoot-like structures are indicated by an arrow; i–l feather-cut leaves of 4-week-old plants incubated for 14 days on medium with the indicated supplements; m 35S::BBM:GR somatic embryos; n immature wild-type zygotic embryo; o somatic embryo formation on wild-type zygotic embryos. MS20 Murashige and Skoog medium with 2% (w/v) sucrose, TDZ thidiazuron, DEX dexamethasone