Development of In Vivo Haploid Inducer Lines for Screening Haploid Immature Embryos in Maize

Doubled haploid technology is widely applied in maize. The haploid inducer lines play critical roles in doubled haploid breeding. We report the development of specialized haploid inducer lines that enhance the purple pigmentation of crossing immature embryos. During the development of haploid inducer lines, two breeding populations derived from the CAU3/S23 and CAU5/S23 were used. Molecular marker-assisted selection for both qhir1 and qhir8 was used from BC1F1 to BC1F4. Evaluation of the candidate individuals in each generation was carried out by pollinating to the tester of ZD958. Individuals with fast and clear pigmentation of the crossing immature embryos, high number of haploids per ear, and high haploid induction rate were considered as candidates. Finally, three new haploid inducer lines (CS1, CS2, and CS3) were developed. The first two (CS1 and CS2) were from the CAU3/S23, with a haploid induction rate of 8.29%–13.25% and 11.54%–15.54%, respectively. Meanwhile, the CS3 was from the CAU5/S23. Its haploid induction rate was 8.14%–12.28%. In comparison with the donor haploid inducer lines, the 24-h purple embryo rates of the newly developed haploid inducer lines were improved by 10%–20%, with a ~90% accuracy for the identification of haploid immature embryos. These new haploid inducer lines will further improve the efficiency of doubled haploid breeding of maize.

Importance of parental genome balance in the generation of novel yet heritable epigenetic and transcriptional states during doubled haploid breeding

BackgroundDoubling the genome contribution of haploid plants has accelerated breeding in most cultivated crop species. Although plant doubled haploids are isogenic in nature, they frequently display unpredictable phenotypes, thus limiting the potential of this technology. Therefore, being able to predict the factors implicated in this phenotypic variability could accelerate the generation of desirable genomic combinations and ultimately plant breeding.ResultsWe use computational analysis to assess the transcriptional and epigenetic dynamics taking place during doubled haploids generation in the genome of Brassica oleracea. We observe that doubled haploid lines display unexpected levels of transcriptional and epigenetic variation, and that this variation is largely due to imbalanced contribution of parental genomes. We reveal that epigenetic modification of transposon-related sequences during DH breeding contributes to the generation of unpredictable yet heritable transcriptional states. Targeted epigenetic manipulation of these elements using dCas9-hsTET3 confirms their role in transcriptional regulation. We have uncovered a hitherto unknown role for parental genome balance in the transcriptional and epigenetic stability of doubled haploids.ConclusionsThis is the first study that demonstrates the importance of parental genome balance in the transcriptional and epigenetic stability of doubled haploids, thus enabling predictive models to improve doubled haploid-assisted plant breeding.

Efficient development of practically usable thermo-photo sensitive genic male sterile lines in wheat through doubled haploid breeding

Two-line hybrid wheat system using thermo-photo sensitive genic male sterility (TPSGMS) is now a dominant and promising approach of wheat heterosis utilization in China. However, few TPSGMS lines available for practical application have always been a bottleneck affecting the efficiency of creating hybrids with strong heterosis since its establishment in 1992. This study aimed to improve the efficiency of developing TPSGMS lines by doubled haploid (DH) breeding. F 1 s and selected F 2 and F 3 sterile plants from 8 crosses made with 2 commercial TPSGMS lines K78S and K456S were used to produce DH lines by wheat × maize system. 24 elite sterile lines possessing stable sterility, good outcrossing and yield potentials, resistance to yellow rust and powdery mildew, and desirable plant height (50-60 cm) were obtained within 4 years. 20 out of 24 elite lines that performed stable sterility in tests of two or three years and high outcrossing rate (>70%) under open pollination, will be used for hybrid breeding later. The percentage of selected sterile lines in total tested DH lines produced from filial generations was in the order of F 3 > F 2 > F 1 for sterility, and F 2 > F 3 > F 1 for elite lines in this study, thus producing DHs from F 2 generation appeared to be the better choice considering the balance of overall breeding efficiency and time saving. This study verified that combining DH techniques with conventional breeding would be an efficient strategy for developing practically usable wheat TPSGMS lines, both in number and time saving.

Influence of different wheat and Imperata cylindrica genetic backgrounds on haploid induction efficiency in wheat doubled haploid breeding

Four Indian and one Japanese accession of Imperata cylindrica were assessed for their influence upon haploid production in F₁ generations of 21 wheat crosses (winter × spring, spring × spring and winter × winter) to find an efficient pollen source for haploid induction, which would enhance doubled haploid breeding in bread wheat. The frequency of haploid induction was influenced differently by the wheat and the I. cylindrica genotypes, indicating both maternal and paternal genetic influence on haploid induction. The gene actions controlling the inheritance of  haploid induction  appeared to be non-additive. Haploid formation efficiency was closely associated with other haploid induction parameters, i.e. pseudoseed formation, embryo formation and haploid regeneration. Amongst wheat F₁ groups, spring × spring wheats exhibited the highest potential for haploid induction. General combining ability for haploid production was highest for the, I. cylindrica genotype Ic-Aru, native to the northeastern Himalayas, which appears as a potential pollen source for efficient haploid induction in bread wheat.