Elsevier

Biotechnology Advances

Volume 33, Issue 6, Part 1, 1 November 2015, Pages 812-829
Biotechnology Advances

Research review paper
Haploids: Constraints and opportunities in plant breeding

https://doi.org/10.1016/j.biotechadv.2015.07.001Get rights and content

Abstract

The discovery of haploids in higher plants led to the use of doubled haploid (DH) technology in plant breeding. This article provides the state of the art on DH technology including the induction and identification of haploids, what factors influence haploid induction, molecular basis of microspore embryogenesis, the genetics underpinnings of haploid induction and its use in plant breeding, particularly to fix traits and unlock genetic variation. Both in vitro and in vivo methods have been used to induce haploids that are thereafter chromosome doubled to produce DH. Various heritable factors contribute to the successful induction of haploids, whose genetics is that of a quantitative trait. Genomic regions associated with in vitro and in vivo DH production were noted in various crops with the aid of DNA markers. It seems that F2 plants are the most suitable for the induction of DH lines than F1 plants. Identifying putative haploids is a key issue in haploid breeding. DH technology in Brassicas and cereals, such as barley, maize, rice, rye and wheat, has been improved and used routinely in cultivar development, while in other food staples such as pulses and root crops the technology has not reached to the stage leading to its application in plant breeding. The centromere-mediated haploid induction system has been used in Arabidopsis, but not yet in crops. Most food staples are derived from genomic resources-rich crops, including those with sequenced reference genomes. The integration of genomic resources with DH technology provides new opportunities for the improving selection methods, maximizing selection gains and accelerate cultivar development. Marker-aided breeding and DH technology have been used to improve host plant resistance in barley, rice, and wheat. Multinational seed companies are using DH technology in large-scale production of inbred lines for further development of hybrid cultivars, particularly in maize. The public sector provides support to national programs or small-medium private seed for the exploitation of DH technology in plant breeding.

Introduction

Sporophytes and gametophytes are the two alternating forms of the life cycle of plants, the former with somatic (2n) and the latter with haploid (n) genomic constitution. Haploid and doubled-haploid (DH) plants have gametophytic (n) and sporophytic (2n) chromosome numbers. Furthermore, a haploid derived from a diploid is known as monoploid, while a haploid derived from a polyploid is a polyhaploid. Haploids occur spontaneously or can be induced by in vivo (inter- and intra-specific hybridization, centromere-mediated haploidization) or in vitro (culture of immature male or female gametophytes) methods (see Section 6). Natural sporophytic haploids in the higher plants were first spotted in Jimson weed (Datura stramonium L.) (Blakeslee et al., 1922), and later noted in several plant species including crops (Chase, 1947, Chase, 1949, Chase, 1969, Chase, 2005, Dunwell, 2010, Maluszynski et al., 2003, Nanda and Chase, 1966). However, doubled haploids were not highly relevant in plant breeding until researchers at the Department of Botany in the University of Delhi, India, reported a breakthrough in the production of haploids from anther culture in Datura (Guha and Maheshwari, 1964, Guha and Maheshwari, 1966), and thereafter through the major discovery of induction of haploids through interspecific crosses followed by embryo culture as a promising method for obtaining haploids in barley (Hordeum vulgare L.) (Kasha and Kao, 1970). Their research revolutionized the use of DH technology in plant breeding worldwide. To date, DH technology has been used in cultivar development in self-fertilizing species, or in inbred line development for their further use in producing hybrids of outcrossing species. Likewise, DH lines (DHLs) derived from hybrid offspring are used as recombinant inbred lines or RILs (Burr et al., 1988) in quantitative genetics research, or for discovering recessive, dominant and deleterious mutations (Castillo et al., 2001, Maluszynski et al., 1996, Szarejko and Forster, 2007 and references therein). DH are used in plant breeding or genetic research because they reach 100% homozygosity after one generation after the induction of haploids (instead of several generations of inbreeding through selfing), the small population size required to obtain a desired genotype (including mutants) from haploids, and last but not the least, the shortening of cultivar development and release.

Globally, DH technology has been effective for developing new cultivars. ‘Maris Haplona’ rapeseed and ‘Mingo’ barley were the earliest releases in Canada (Ho and Jones, 1980, Thompson, 1972), while most recent releases in wheat were ‘BRS 328’ in Brazil and ‘Emerson’ in Canada (Graf et al., 2013, Scheeren et al., 2014) or ‘Kharoba’ in Morocco (Elhaddoury et al., 2012). In excess of 300 DH-derived cultivars, with more than 100 cultivars each in barley and rice and above 50 rapeseed cultivars were reported (Chen, 1986, Daofen, 1986, De Buyser et al., 1987, DePauw et al., 2011, Dunwell, 2010, Elhaddoury et al., 2012, Forster and Thomas, 2005, Forster et al., 2007, Graf et al., 2003, Graf et al., 2013, Hu and Zeng, 1984, Humphreys et al., 2006, Humphreys et al., 2007, Humphreys et al., 2013, Jain et al., 1996, Kang et al., 2011, Loo and Xu, 1986, Palmer et al., 2005, Pauk et al., 2009, Sadasivaiah et al., 2004, Sãulescu et al., 2012, Scheeren et al., 2014, Thomas et al., 2003, Tuvesson et al., 2007, Yang and Fu, 1989, Zhao et al., 1990, Zhu and Pan, 1990). DH-derived cultivars currently occupy significant acreage in some countries. For example, 25 wheat cultivars accounted for more than one third of the Canadian wheat acreage, with Lillian and AC Andrew being the most widely grown wheat cultivars in Canada (DePauw et al., 2011), or a DH-derived wheat cultivar Glossa grew in 16% of the total wheat area (300,000 ha) just in 5 years after its release in Romania (Sãulescu et al., 2012). The Peruvian highland barley farmers benefitted the most by growing DH-derived barley lines (Ya/LM94-PC27, B12/LM94-PC34), while the researchers in Peru saved 26% research cost by adopting DH technology in barley breeding program (Gomez-Pando et al., 2009).

In vivo induction of DH is widely adopted method for inbred line development in maize (Zea mays L.) (Geiger and Gordillo, 2009, Prasanna et al., 2012). Unlike barley, maize, oat (Avena sativa L.), rice (Oryza sativa L.), rye (Secale cereale L.), and wheat among cereals (Forster et al., 2007, Jauhar et al., 2009, Germanà, 2011, Prasanna et al., 2012, Tadesse et al., 2012, Niu et al., 2014; http://www.agriculture.gov.sk.ca/agv1309-pg12), Brassica species among oilseeds (Xu et al., 2007) and potato (Solanum tuberosum L.) among tuber crops (Rokka, 2009), the DH technology in other crops including legumes (Croser et al., 2006) and root crops (Perera et al., 2014) has not reached to the stage leading to its use in plant breeding. In recent years a technology-driven approach such as centromere-mediated genome elimination procedure for the development of DH, initially proposed in Arabidopsis (Comai, 2014, Ravi and Chan, 2010), have been undertaken in banana (Musa spp.), barley, Brachypodium, cassava (Manihot esculenta Crantz), Gossypium, Lotus japonicus, rice (Oryza sativa L.), soybean (Glycine max (L.) Merr.), sugarbeet (Beta vulgaris L.), switchgrass (Panicum virgatum L.), and tobacco (Nicotiana tabacum L.) (Tek et al., 2014). Although a few years have passed since the technique's development in Arabidopsis, there have been no published successes in other plant species.

Although the development of new cultivars is urgently needed to meet the demands of an increasing population and the challenges of a changing climate, cultivar development is a lengthy and time-consuming process. New methods that enhance the efficiency of plant breeding are under investigation. Today, most crops have abundant genomic resources (Dwivedi et al., 2007), high throughput cost-effective phenotyping (Araus and Cairns, 2014, Cobb et al., 2013, Fiorani and Schurr, 2013), and genotyping tools (Thudi et al., 2012, Varshney et al., 2009). Likewise, the information on markers and genomic regions associated with agronomically beneficial traits (Dwivedi et al., 2007, Thudi et al., 2012, Varshney et al., 2009, Varshney et al., 2013), and the genome sequences of many food crops (Bevan and Uauy, 2013, Hamilton and Buell, 2012) offer knowledge that has been used to breed new cultivars (Collard and Mackill, 2008, Dwivedi et al., 2007, Mba et al., 2012, Poland and Rife, 2012, Varshney et al., 2013, Varshney et al., 2014). Above all, the scientific knowledge generated through DH technology has been enhanced in some crops such as barley, Brassica spp., maize, rice, triticale (xTriticosecale Wittm.) and wheat, and should be integrated with phenomics and genomics to accelerate cultivar development and economize plant breeding operations. This article deals with the induction and identification of haploids, factors influencing haploid induction, in vitro manipulation of gametic tissues for plant breeding, molecular basis of microspore embryogenesis, agrobacterium-mediated genetic transformation to support androgenesis, centromere-mediated genome elimination for induction of haploids, the genetics of haploid induction and breeding efficiency, trait fixation and unlocking new genetic variation from landraces, and the establishment of state of the art technology to support plant breeding programs for DH induction.

Section snippets

Factors affecting gametophytic haploid production

The genotype of the donor plant determines the efficiency of in vitro (Chen et al., 2011, Datta, 2005, Nitsch and Nitsch, 1969) or in vivo (Bitsch et al., 1998, Garcia-Llamas et al., 2004) haploid production. This response varies not only among species but also within a species, with few genotypes having great response while others being recalcitrant. For example, Brassica napus is more responsive to microspore embryogenesis compared to Brassica juncea, and winter genotypes are more responsive

Gametoclonal variation

The variation observed among plants regenerated from cultured gametic cells is termed gametoclonal variation (Evans et al., 1984), for example, variation for several agronomic traits was noted in wheat (but not in barley) using bulbosum method of DH production (Snape et al., 1988). Variation from gynogenically derived tef (Eragrostis teff (Zuccagni)), an important cereal crop of Ethiopia, was found for plant height, panicle length, culm thickness, seed size, and maturity (Gugsa and Loerz, 2013,

Molecular basis of microspore embryogenesis

Microspore embryogenesis involves reprogramming of the pollen immature cell towards embryogenesis. The use of functional genomic tools has allowed the identification of genes associated with microspore embryogenesis (ME) in barley and rapeseed (Joosen et al., 2007, Malik et al., 2007, Maraschin et al., 2006, Muñoz-Amatriaín et al., 2006, Muñoz-Amatriaín et al., 2009, Seguí-Simarro and Nuez, 2008, Tsuwamoto et al., 2007). There are 14 genes (TaTPD1-like, TAA1b, GSTF2, GSTA2, TaNF-YA, TaAGL14,

Identifying putative haploids (focusing on recent advances in maize)

A key issue for the commercialization of DH technology is the development of an efficient system for identification of putative haploids. This becomes crucial when using in vivo DH technology for induction and identification of haploids from hybrid seeds. Several methods are now available to identify haploids from hybrids as discussed herewith.

Conventional methods to induce doubled haploids

The in vitro procedure using androgenesis (anther or microspore culture) and gynogenesis (unfertilized egg cell) has been used to produce DH. Androgenesis refers to culturing immature anther or microspores from the immature pollen grain in artificial media to isolate haploid cells that are then chromosome doubled using colchicine to develop DH. It is beyond the scope of this review to provide a depth discussion on DH production using this technology because there are excellent articles on this

Agrobacterium-mediated genetic transformation to androgenesis

Using a novel genetic transformation method based on infection of androgenic pollen cultures with Agrobacterium tumefancies, Kumlehn et al. (2006) produced primary transgenic (T0) plants in barley carrying a single copy of the sequence integrated, of which about 60% of the plants set seed, indicating spontaneous genome doubling, which opens up the opportunity for production of doubled haploid T1 seeds instantly homozygous for the transgene. More importantly, this method has great potential for

Centromere-mediated genetic engineering to haploid induction

As discussed above, haploid plants can be induced through the regeneration of plants from haploid (gametic) tissues, through the destruction of a single parent's genome (for example, through irradiation of pollen) or through the selective loss of one parent's chromosomes during early embryogenesis (Sanei et al., 2011) (which can be induced by interspecific hybridization or by “haploid inducing” variants). All of the above techniques work for some species, or cultivars, but not for others. For

Insights into the genetics of haploid induction

The donor plant genotype affects haploid induction and the subsequent embryo regeneration, as noted by their induction rates, in maize (Röber et al., 2005), onion (Allium cepa L.) (Alan et al., 2004, Bohanec and Jakse, 1999, Gioffriau et al., 1997), summer squash (Cucurbita pepo L.) (Shalaby, 2007) or sweetpotato (Ipomea batata (L.) Lam) (Kobayashi et al., 1993), which highlights the genotypic-specific response to gynogenesis. This maternal in vivo haploid induction ability in maize is a

Trait fixation (heterosis) via anther culture

The economics of heterosis is limited by costs of hybrid seed production, and farmers purchasing these seeds every season to realize the yield potential of hybrid cultivars. The DH technique provides unique opportunity to develop lines that are truly homozygous and have excellent yields approaching hybrid cultivars. This proof of concept was demonstrated when some DH lines developed via anther culture from the heterotic F1 crosses achieve the yield of heterotic hybrids in mutant crosses of

Unlocking new genetic variation

Landraces are valuable plant genetic resources, which evolved over time and adapted to the natural environments, with high capacity to tolerate stress. They also yield reasonably well under low input production systems. The landraces are highly heterogeneous genepool (which could show some undesirable “genetic load”) and may be excellent resources to identify new sources of variation associated with agronomically beneficial traits for use in plant breeding. Inbreeding unveils any genetic load,

Accelerating crop breeding using doubled haploids, DNA markers, and data management

The development of crop cultivars by crossbreeding is both time and resource consuming. It takes between 8 to 10 years from the time the cross is made until phenotypically advanced uniform lines are produced. These are then evaluated for at least 3 years to identify potential candidate lines for cultivar release. The continued demand for new cultivars with specific characteristics requires that adopted plant breeding methods accelerate the development of the new cultivars. An off-season nursery

Genetic gains through haploid breeding vis-à-vis other crossbreeding methods

Genetic gains (ΔG) depend on additive genetic (σ2A) and phenotypic variances (σ2P), which are used to estimate a ratio known as narrow sense heritability (h2 = σ2A2P), selection intensity (or the percentage of individuals selected and advanced to the next generation), parental control of males and females (c), and time. The plant breeding equation for ΔG per cycle is (ΔC) = Κ c h2 σP, where Κ is the selection differential in standard deviation units. The genetic gain per year (ΔG/Y) is more

Perspectives

Ninety years after the discovery of natural sporophytic haploids in Jimson weed and about 50 years after the breakthrough on the production of haploids from anther culture in Datura and the use of bulbosum method for haploid induction in barley, haploids and DHs are broadly used in breeding various crops. DH research has advanced considerably and facilitated the release of large number of cultivars, mostly in Brassica and cereals. Research led to great understanding of the genetics and

Acknowledgment

We appreciate the four anonymous reviewers for making useful suggestions on improving the manuscript. Sangam Dwivedi highly appreciates Ms Ishrath Durafsha of Knowledge Sharing and Innovation Program of ICRISAT for arranging reprints.

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      Doubled haploid (DH) wheat plants are generated where the gamete set of chromosomes (n) has been doubled to establish a balanced hexaploid genome. The genomes of such plants are completely homozygous, and they are of great importance in breeding acceleration (Fig. 1) as well as genetics, genomics, genetic transformation and gene editing (Hussain et al., 2012; Dwivedi et al., 2015; Rustgi et al., 2017; Tadesse et al., 2019; Budhagatapalli et al., 2020; Ferrie et al., 2020). The first example of spontaneous haploidy was shown for Datura stramonium L. (Jimson weed) and this was published a hundred years ago (Blakeslee et al., 1922).

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