Methods for characterizing pollen fitness in Cannabis sativa L.

Abstract

Pollen grains are male gametophytes, an ephemeral haploid generation of plants, that commonly engage in competition for a limited supply of ovules. Since variation in reproductive capabilities among male gametophytes may influence the direction and pace of evolution in populations, we must be able to quantify the relative fitness of gametophytes from different sires. To explore this, we estimated the relative fitness of groups of male gametophytes in a dioecious, wind-pollinated model system, Cannabis sativa, by characterizing the non-abortion rate (measured via chemical staining) and viability (measured via in vitro germination) of pollen from multiple sires. Pollen viability quickly declined within two weeks of anther dehiscence, and pollen stored under freezer conditions did not germinate regardless of storage time. In contrast, pollen non-abortion rates declined slowly and persisted longer than the lifetime of a sporophyte plant under both room temperature and freezer conditions. Pollen samples that underwent both viability and non-abortion rate analysis displayed no significant correlation, implying that researchers cannot predict pollen viability from non-abortion rates, nor infer male gametophytic fitness from a single measure. Our work demonstrates two independent, differential approaches to measure proxies of male fitness in C. sativa.

Can They Keep Up? Fecundity, Pollen Fitness, and Reproductive Success in Subdioecious, Sexually Labile Cannabis Sativa L

“…My second empirical chapter (Chpt. 3;Wizenberg et al, 2022) validated two different methods of quantifying relative pollen fitness and demonstrated no relationship between these two fitness estimates -justifying our choice to use both as indicators of male reproductive performance.…”

“…The experimental design, data collection, and analysis was a joint effort between Dr. Campbell and I. Michelle provided assistance throughout, aiding in the preparation of materials, troubleshooting, and contributing to preparation of the final manuscript. This manuscript has been published with the following citation: Wizenberg, S. B., Dang, M., and Campbell, L. G. (2022). Methods for characterizing pollen fitness in Cannabis sativa L. PloS One, 17(7), e0270799.…”

“…1). Using methods we previously developed for measuring pollen abundance (Wizenberg et al, 2020) and pollen fitness (Wizenberg et al, 2022), we asked:…”

Discovery of A Novel Hemp Branching Mutant

“…Cannabis can be both monoecious and dioecious (Chandra et al 2020). Additionally, its pollen can be dispersed up to 5 km downwind and remain viable for long periods, resulting in diverse forms of natural cross-breeding between cultivars or varieties (Small & Antle, 2008; Wizenberg et al 2022). This helps to explain the high heterozygosity of early cannabis genetic resources.…”

Environmental metagenetics unveil novel plant‐pollinator interactions

“…Due to their sessile nature, plants rely on external vectors for dispersal of gametophytes; they are incapable of physically finding a mate, so they depend on abiotic or biotic forces to facilitate the fusion of gametes. Their intergenerational life cycle results in ephemeral haploid gametophytes that act as links between parental plants and their offspring – dispersal of these specialized cells is essential for reproductive success (Robledo‐Arnuncio et al., 2014 ; Snow & Lewis, 1993 ; Wizenberg et al., 2022 ). Most plant species rely on either anemophily (wind pollination) or entomophily (insect pollination) as a dispersal mechanism, frequently evolving specialized pollen morphology to maximize success under one of those two conditions (Lu et al., 2022 ; Pacini & Franchi, 2020 ; Wizenberg et al., 2020 ).…”