Identification of maternal haploids of Nicotiana tabacum aided by transgenic expression of green fluorescent protein: evidence for chromosome elimination in the N. tabacum × N. africana interspecific cross

Abstract

The Nicotiana tabacum 9 N. africana cross is a semilethal cross, where greater than 99 % of progeny die in the cotyledonary stage due to an interspecific lethality reaction. The low frequencies of surviving plants are mixtures of maternal tobacco haploids and aneuploid F 1 hybrids. For effective use of surviving haploids in a breeding program, an efficient method is needed to distinguish them from aneuploid hybrids during the seedling stage. The first objective of this research was to investigate the use of N. africana engineered with a green fluorescent protein (gfp) transgene for distinguishing haploids from other surviving plants derived from this cross. Results demonstrated gfp expression to be a useful phenotypic marker for separation of aneuploid hybrids at the seedling stage. Microsatellite marker genotyping, flow cytometry determinations, and chromosome counts resulted in identification of progeny that exhibited intermediate DNA amounts and variable chromosome numbers donated by the N. africana parent. Three individuals with near-haploid DNA content were identified that carried N. africana markers and n = 24 ? 2 or n = 24 ? 3 chromosomes. The results present evidence of genetic instability in progeny from the N. tabacum 9 N. africana cross and suggest that, in contrast to previous belief, chromosome elimination may play at least a partial role in the generation of haploids from this cross. Additionally, microsatellite marker genotyping demonstrated a role for a gene or genes located near the end of N. tabacum chromosome H (linkage group 11) in the interspecific lethality reaction.

Overcoming Hybrid Lethality Induced by Chromosomal Instability in an Interspecific Hybrid of Genus Nicotiana

“…Speci cally, 11 hybrid seedlings that had overcome lethality were obtained, and the frequency of appearance of hybrid seedlings that overcame lethality in this cross combination was calculated to be 8.5 x 10 -4 . This value was close to 1.1 x 10 -3 , which is the frequency of appearance of hybrid seedlings that overcame lethality in a previous study of the N. tabacum x N. africana cross 18 .…”

“…Additionally, Ma 11 observed that the H chromosome belongs to the S genome. Therefore, we infer that the H chromosome in the reports of Hancock et al 18 and Ma 11 is the same as the Q chromosome in the report of Tezuka et al 17 . If this interpretation is valid, we consider it highly likely that Nt6549g30 is involved in the lethality of the N. suaveolens x N. tabacum cross.…”

“…On the other hand, the SSR marker of Linkage group No. 11, a gene that is detected on the Q chromosome of the S genome 17 , also was detected on the H chromosome 18 . Additionally, Ma 11 observed that the H chromosome belongs to the S genome.…”

“…In some lethal cross combinations between N. tabacum and wild Nicotiana species, including N. suaveolens x N. tabacum, viable hybrid seedlings (i.e., hybrid seedlings that overcome lethality) appear spontaneously at a certain frequency. Hancock et al 18 detected hybrid seedlings that overcame lethality at a frequency of about 1.1 x 10 -3 in a N. tabacum x N. africana cross. Those authors also con rmed that the SSR marker at the distal part of Linkage group No.…”

Introgression Analysis and Morphological Characterization of an Arachis hypogaea × A. diogoi Interspecific Hybrid Derived Population

“…The unique genomic composition of two different parental genomes within one nucleus can lead to intergenomic conflicts such as chromosome elimination. Chromosome elimination has been reported in interspecific hybrids within other crop species genera such as Hordeum (Subrahmanyam and Kasha, 1973) and Nicotiana (Hancock et al, 2015). Hypotheses to explain the mechanisms of chromosome elimination after interspecific hybridization in various crop species include differences in timing of the mitotic cycle between the two species, histone changes resulting in the failure of centromeres to attach to the spindle microtubules, host‐specific nucleases capable of degrading alien chromatin, or the formation of micronuclei followed by selective removal (Kasha and Kao, 1970; Subrahmanyam and Kasha, 1973; Finch, 1983; Gernand et al, 2005; Ravi and Chan, 2010, 2013).…”