Optimized Protocol for Development of Androgenic Haploids and Doubled Haploids in FCV Tobacco (Nicotiana tabacum)

Abstract

Haploids are plants with gametophytic chromosome number, which upon chromosome duplication results in production of doubled haploids (DHs). There are several methods to obtain haploids and DHs, of which in vitro anther culture is the most effective and widely used method in tobacco. The production of haploids and DHs through androgenesis allows for a single-step development of complete homozygous lines from heterozygous genotypes, shortening the time required to produce homozygous genotypes in comparison to the conventional breeding scheme. The DH development process comprises two main steps: induction of androgenesis and duplication of the haploid genome. The critical stages of DH protocol in tobacco are determining the bud stage for anther culture, pretreatment, anther culture media, detection and identification of haploids, and chromosome doubling. Here we present an efficient anther culture protocol to get haploids and DHs in flue-cured virginia (FCV) tobacco. This optimized protocol can be used as a potential tool for generation of haploids and DHs for genetic improvement of tobacco.