Crop wild relatives may hold the genetic tools needed to help agriculture withstand drought, heat, salinity, emerging diseases and increasingly unstable growing conditions. Yet the same evolutionary distance that preserved these valuable traits often prevents breeders from moving them into modern crops. A new review in Theoretical and Applied Genetics describes how “cross incompatibility”—a collection of reproductive barriers that block fertilization, embryo development or chromosome pairing—restricts the exchange of genomes between cultivated plants and their wild relatives. The authors argue that advances in genomics, genome editing, embryo culture, synthetic biology and artificial pollination could transform these barriers from fixed biological obstacles into traits that breeders can predict, manipulate and, in some cases, redesign.
The problem begins long before a hybrid seed is formed. In flowering plants, pollen must first land on a receptive stigma, hydrate, germinate and produce a pollen tube capable of growing through the style toward the ovule. At every stage, molecular recognition systems compare pollen with pistil tissues. If the pollen is rejected, germination may fail; if the tube grows too slowly or encounters an incompatible biochemical environment, it may stop before reaching the female gametophyte. These pre-zygotic barriers are particularly important in wide crosses involving different species or genera. In some tomato crosses, for example, pollen from the cultivated species is arrested inside the style of a wild relative. In peppers, pollen may fail to germinate on the stigma of another species. Such failures prevent fertilization even when both parents are otherwise fertile.
Successful pollen-tube growth does not guarantee a viable hybrid. After fertilization, the embryo and the surrounding endosperm must develop according to a tightly coordinated genetic and genomic balance. The endosperm, a nutritive tissue formed through double fertilization, is especially sensitive to differences in parental genome dosage and imprinting. When related plants with different chromosome numbers are crossed, the endosperm may collapse, starving the embryo and causing the seed to abort. This phenomenon, often called the triploid block, is a major obstacle in crosses between plants of different ploidy levels. Even when an embryo survives, chromosomes from the two parents may fail to pair correctly during meiosis. The resulting plants can produce little or no viable pollen or eggs, making further breeding difficult.
These barriers are not controlled by a single universal “incompatibility gene.” They range from pollen–pistil recognition systems and cell-wall chemistry to embryo development, chromosome behavior and epigenetic regulation. In maize, the Gametophyte factor1, or Ga1, system is one of the clearest examples of a genetically controlled pre-fertilization barrier. Linked genes at the locus influence both pollen performance and the silk tissues through which pollen tubes must grow. Pectin methylesterases, enzymes that modify the cell wall, help determine whether a pollen tube can navigate the silk. Related systems involving Ga2 and Tcb1 create additional compatibility patterns. Depending on the parental combination, pollen may be accepted, slowed or completely rejected, producing unilateral incompatibility in which one crossing direction succeeds while the reciprocal cross fails.
Rice provides a different illustration of how reproductive barriers can arise after fertilization or during gamete formation. Hybrid sterility between the indica and japonica subspecies is associated with loci including S5, Sa and DPL1/DPL2. At S5, alternative alleles can affect embryo-sac development and determine whether hybrids are fertile. Other loci influence pollen viability or cause selective loss of gametes carrying particular genetic combinations. These systems demonstrate that incompatibility may result from interactions between genes inherited from different evolutionary backgrounds rather than from a simple mismatch at one location. In wheat, crossability genes such as Kr1 and Kr2 restrict hybridization with related species, while in Brassica, the S-locus uses receptor–ligand signaling between stigma and pollen to control recognition. The review emphasizes that identifying these mechanisms is essential because the best strategy depends on where the cross fails.
Conventional breeding has already produced striking successes by bypassing, rather than removing, reproductive barriers. Embryo rescue is among the most widely used methods. Breeders pollinate the plants, wait until a young embryo forms, then remove it before the surrounding endosperm deteriorates and culture it on a nutrient medium. The rescued embryo can develop into a plant that would otherwise never survive as a seed. Ovule, ovary and siliqua culture apply related principles at slightly different developmental stages. Bridge crosses offer another route: a difficult cross is performed in stages through a compatible intermediate species. In tomato, breeders used Lycopersicon chilense as a bridge to transfer traits from the incompatible wild species L. peruvianum into cultivated tomato. Somatic hybridization takes the process further by fusing protoplasts—plant cells stripped of their walls—from different species or genera, allowing their genomes to combine without relying on normal pollen–pistil interactions.
Breeders have also experimented with the reproductive environment itself. Sugar solutions can improve pollen germination and pollen-tube growth, while salt treatments, hormones and specialized pollination methods have helped overcome pre-fertilization barriers in crops such as papaya, lily and chrysanthemum. Mentor pollen, delayed pollination and treatments with auxins, cytokinins or gibberellic acid can alter the physiological conditions around the stigma and style, sometimes giving otherwise weak pollen enough time to reach the ovule. These methods are often empirical and highly crop-specific, but they remain valuable because they can be applied without permanently altering the genome. Polyploidization, meanwhile, can restore fertility by providing additional homologous chromosomes for pairing or by stabilizing hybrid genomes, although it may also create new incompatibilities and developmental abnormalities.
Modern genomics is making these interventions more precise. A single reference genome can miss structural variants, duplicated genes, transposable-element insertions and entire stretches of DNA that differ among cultivated lines and wild relatives. Pan-genomes, and especially graph-based pan-genomes, represent these alternate genomic configurations more accurately. For incompatibility research, this matters because reproductive loci are often complex, rapidly evolving or organized in tightly linked gene clusters. A graph could reveal structural differences around S-loci, pollen-expressed genes, stylar recognition factors, small-RNA regions and endosperm regulators that would be invisible in a conventional linear alignment. The authors propose building compatibility-focused pan-genomes containing compatible and incompatible parents, wild donors, bridge species and derived hybrids. Such resources could enable “compatibility haplotype mining,” allowing breeders to identify parental combinations likely to support pollen growth, fertilization and embryo survival before investing in large crossing experiments.
The emerging picture also extends beyond DNA sequence. Transposable elements can reactivate when divergent genomes meet, disrupting gene regulation and contributing to structural variation, chromosome rearrangements and hybrid defects. In citrus, a miniature inverted-repeat transposable element inserted into the promoter of an S-RNase gene has been linked to the loss of self-incompatibility, showing how mobile DNA can alter reproductive behavior. Small RNAs, DNA methylation and genomic imprinting also influence whether hybrid endosperm develops normally. In Arabidopsis, chemically induced epimutagenesis using the DNA-methyltransferase inhibitor 5-azacytidine has been reported to bypass some reproductive barriers. These findings suggest that compatibility can depend not only on which genes are present, but also on when they are active, how they are silenced and how parental genomes are balanced inside developing seeds. Single-cell transcriptomics, pollen RNA profiling, methylome analysis and embryo–endosperm sequencing could help breeders pinpoint the exact developmental moment when a cross fails.
Genome editing could then turn that knowledge into targeted intervention. CRISPR-based disruption of incompatibility genes has already been used to modify reproductive barriers in several crops. In maize, editing a female component of the Ga1 system has enabled crosses that would otherwise be blocked. In potato, simultaneous editing of S-RNase and HT genes has increased seed production by weakening self-incompatibility pathways. Related approaches have been explored in tomato and rapeseed. More refined tools, including base editors and prime editors, could alter individual nucleotides or regulatory sequences without creating large DNA breaks. The goal would not always be to eliminate incompatibility permanently. A breeder might instead create a line that is compatible during a controlled cross but retains reproductive containment in the field. Synthetic biology could eventually add inducible receptors, engineered pollen–stigma signals or molecular switches that activate or suppress pollen rejection at selected times. Such systems could make compatibility programmable, although their ecological and regulatory risks would require careful assessment.
The review also highlights a less obvious possibility: using insects, robotics and artificial intelligence to improve the physical delivery of pollen. Managed bees and other pollinators can transfer pollen across flowers with greater precision than hand pollination and may help overcome differences in floral structure, flowering time or pollen placement. Recent work has reported successful intergeneric hybrids in Brassica rapa and Sinapis alba using insect-mediated pollination. Computer vision systems can monitor flower opening, stigma receptivity and pollinator visits, while robotic platforms can locate flowers and deliver pollen to selected targets. These tools cannot repair a lethal genetic incompatibility, but they may solve logistical barriers that are often mistaken for biological ones. Combined with molecular pollen tagging, they could reveal which insects move compatible pollen, when pollen transfer is most effective and which reciprocal crossing direction has the greatest chance of success. Machine-learning models could integrate parental genomes, chromosome numbers, flowering synchrony, pollen viability, structural variation and embryo-rescue results to predict whether a proposed cross is likely to be compatible.
The stakes are high because many traits needed for climate-resilient agriculture remain concentrated in crop wild relatives and in the secondary or tertiary gene pools. These plants may carry resistance to pathogens, tolerance to drought or salinity, and adaptations to poor soils or extreme temperatures, but their value cannot be realized if their genomes remain inaccessible. The authors caution that major gaps remain: many candidate genes have not been functionally validated, predictions have rarely been tested across field environments, and compatibility mechanisms in a small group of model crops cannot automatically be transferred to other species. Engineering reproductive compatibility could also increase unintended gene flow into wild or weedy relatives. Even so, the convergence of pan-genomics, multi-omics, genome editing, embryo rescue, synthetic biology and precision pollination offers a new framework. Instead of treating cross incompatibility as an impenetrable wall, breeders may increasingly view it as a complex, measurable system—one that can be mapped, forecast and selectively reprogrammed to expand the genetic foundation of future crops.
Subject of Research: Cross incompatibility and reproductive barriers restricting genome exchange between cultivated plants and crop wild relatives.
Article Title: Cross incompatibility restricting genome exchange and diversification of primary gene pool in plants: challenges and opportunities
Article References: Thudi M, Naik YD, Jha UC et al. Theoretical and Applied Genetics 139, article 245 (2026). https://doi.org/10.1007/s00122-026-05347-x
Image Credits: AI Generated
DOI: https://doi.org/10.1007/s00122-026-05347-x
Keywords: Cross incompatibility, crop wild relatives, plant breeding, reproductive barriers, embryo rescue, somatic hybridization, genome editing, CRISPR, pan-genomics, hybrid sterility, pollen–pistil interactions, polyploidy, synthetic biology, precision pollination, crop resilience
Tags: advances in plant genomics and gene editingartificial pollination techniques for genome transferembryo culture and synthetic biology in crop breedinggenetic tools for managing cross incompatibilitygenome exchange barriers in cropsimpact of reproductive barriers on crop improvementmolecular mechanisms of pre-zygotic barriersovercoming reproductive incompatibility in agricultureplant cross-incompatibilityreproductive barriers in plant breedingrole of molecular recognition in plant fertilizationwild-relative gene pool diversification
