Development and validation of CAPS-marker associated with the Rf2 gene in sorghum (Sorghum bicolor (L.) Moench)

Background. The development of heterotic hybrids based on cytoplasmic male sterility (CMS) is the leading strategy in breeding sorghum (Sorghum bicolor (L.) Moench). The trait of pollen fertility restoration in forms with CMS A1 (milo), predominantly used in sorghum breeding, is determined by at least two dominant complementary genes Rf1 and Rf2, and also gene Rf5. The development of accessible molecular markers of sorghum Rf genes is highly relevant for hybrid breeding, since they can significantly accelerate the process of creating female sterile forms (A lines), sterility maintainers (B lines) and pollen fertility restorers (R lines).Material and methods. The studied material included 36 sorghum accessions from the VIR collection, which differed by the ability to restore pollen fertility in forms with A1-type CMS. The nucleotide polymorphism of 935 bp fragments of the PPR genes Sobic.002G057050, Sobic.002G054100, and Sobic.002G054200 located at the chromosome 2 was studied.Results. The fragments obtained with the use of a pair of 2459403fw and 2459403 primers were 935 bp long and included parts of three genes: Sobic.002G057050, Sobic.002G054100, Sobic.002G054200. For identifying the sequence variant Sobic.002G057050-1090 associated with the Rf2 gene, Tru9 I restrictase was chosen, which allows obtaining a 572 bp fragment unique for all the studied R lines. Such a marker was found in 10 sorghum lines from West China and Kyrgyzstan, which are widely used in breeding as fertility restorers. The fragment was found neither in three lines with sterile cytoplasm and their fertile analogues, nor in 7 accessions of kafir sorghum, which lacked functional alleles of Rf genes.Conclusions. It has been demonstrated that the marker can be used for selection and checking purity of R and B/A lines. It is also applicable for verifying hybridity of F1 seeds and analyzing hybrid populations from crosses of R lines 924-4, 928-1, 929-3, 931-1, 933-1/6, 1237-3, 1243-2, 1251, 1150-1, F10BC2 with A lines Nizkorosloe 81s, А-83 and А-10598. It may be suggested that the ability to restore pollen fertility in R lines, which lack the marker CAPS- 572, is determined by another Rf gene. The studied 935 bp fragment of Sobic.002G057050 harbours 22 SNP, therefore the development of CAPS-markers for their identification and differentiation can be promising.

Variation of morphological indicators of three-line hybrids sunflower and their parental forms

The article presents the results of three-year (2016–2018) studies of five three-line hybrids of the IMC selection - Agent, Agronomichny, Marshal, Kamenyar, Zaporozhskij 28 and their parent components - simple unreduced hybrids - ZL22A/102B, ZL42A/46B, ZL42A/58B and pollen fertility restorers – ZL512V, ZL678V and ZL7034V. It was found that individual indicators do not change synchronously. The Agent, Marshal and Kamenyar hybrids had the highest plant height in 2016, one each in 2017 – Agronomichny and in 2018 – Zaporozhskij 28. At the same time, three hybrids (Agent, Agronomical, Marshal) had the largest basket diameter in 2018 and two (Zaporozhskij 28, Kamenyar) in 2017. The shortest growing season hybrids Marshal, Zaporozhskij 28 and Kamenyar had in 2016 (90, 105 and 105 days), and two – Agent and Agronomichny in 2018 – 100 and 103 days, respectively. Among the simple unrecovered hybrids, two – ZL42A/46B and ZL42A/58B had high indicators of plant height, basket diameter and duration of the growing season in 2016. In the ZL22A/102B hybrid, the diameter of the basket was also the largest in 2016 (18.5 cm), the average plant height in 2016 – 124.5 cm was slightly lower than in 2018 (125.4 cm), also in these years in it almost coincided with the duration of the growing season – 97 days in 2016 and 96 in 2018 Fertility restorers had the highest indicators of plant height and basket diameter in 2017, also this year they had the shortest growing season, in 2016, on the contrary, they had the lowest height and the diameter of the basket, and the long growing season. The size of the baskets in the Marshal and Kamenyar hybrids correlates with the moisture supply of plants and the hydrothermal coefficient – the correlation coefficients are 0.997, 0.902 and 0.990, 0.973, respectively. The phases of organogenesis of plants of hybrids Kamenyar and Zaporozhskij 28 are greatly influenced by temperature, the correlation coefficient of this indicator with plant height and with the duration of the growing season is 0.996 for Kamenyar and 0.946 for Zaporozhskij 28, and with a hydrothermal coefficient – 0.939 and 0.753. In the Agent hybrid, the temperatures in June have the greatest influence on plant growth - the correlation coefficient is 0.997 and precipitation in May is 0.968, and the temperatures in May - 0.999 and June - 0.998 on the size of the basket. For the height of plants and the duration of the growing season at Agronomichny, moisture availability at the beginning of the growing season is very important, the correlation coefficients are 0.918 and 0.994, and in August during the filling of seeds 0.996 and 0.927, as well as July precipitation is 0.995. Of the simple unrecovered hybrids, the most demanding for heat is ZL22A/102B - the correlation coefficients are 0.941 with plant height and vegetation duration, 0.843 with the basket size. For ZL42A/58B, the most important are the May precipitation, their correlation with the vegetation duration of 1,000 and the basket diameter of 0.987, and the July temperatures - 0.999 and 0.993, respectively. Among the pollen fertility restorers, the line ZL512V turned out to be the most demanding to weather conditions. For plant growth, the temperatures of April (0.906), May (0.995) and June (1.000) are very important, for the duration of the growing season respectively – 0.958, 0.971 and 0.991, and for the size of the basket, precipitation in May (0.956) and July temperatures (0.943). The correlation coefficient with the sum of active temperatures is – 0.829 for plant height – 0.851 for basket diameter – 0.902 for the growing season.

The radish Ogura fertility restorer impedes translation elongation along its cognate CMS-causing mRNA

The control of messenger RNA (mRNA) translation has been increasingly recognized as a key regulatory step for gene control, but clear examples in eukaryotes are still scarce. Nucleo-cytoplasmic male sterilities (CMS) represent ideal genetic models to dissect genetic interactions between the mitochondria and the nucleus in plants. This trait is determined by specific mitochondrial genes and is associated with a pollen sterility phenotype that can be suppressed by nuclear genes known as restorer-of-fertility (Rf). In this study, we focused on the Ogura CMS system in rapeseed and showed that reversion to male sterility by the PPR-B fertility restorer (also called Rfo) occurs through a specific translation inhibition of the mitochondria-encoded CMS-causing mRNA orf138. We also demonstrate that PPR-B binds within the coding sequence of orf138 and acts as a ribosome blocker to specifically impede translation elongation along the orf138 mRNA. Rfo is the first recognized fertility restorer shown to act this way. These observations will certainly facilitate the development of synthetic fertility restorers for CMS systems in which efficient natural Rfs are lacking.

Phenotypic Characterization of Diverse Rice Fertility Restorers

Characterization of restorer lines of rice based on seedling and plant morphology contributes in assessment of varietal identity and further its utilization in development of hybrids. With this objective the work was carried over with 90 restorer lines with three replications during Kharif season 2018. The experiment conducted under Rice Improvement Project at Seed Breeding Farm, Jawaharlal Nehru Krishi Vishwa Vidyalaya, Jabalpur (M.P.).Out of 41 morphological traits as per DUS guidelines, variability was reported for all traits except leaf sheath anthocyanin colouration, presence of leaf auricle, presence of leaf collar, presence of leaf ligules, white colour of leaf ligules, absence of stem anthocyanin colouration of internodes, male sterility and presence of secondary branching in panicle. The genotypes Mahamaya, ANP-526 and ANP-553 revealed distinctness for most of the traits.

Promising experimental hybrids of spring rapeseed (Brassica napus L.) for CMS-system of Polima type

One of the prior directions in spring rapeseed breeding is development of heterotic hybrids. Seed growing of the most hybrids F1 used in production are often carried out by two types of cytoplasmic male sterility, they are polima and ogura. At the Lipetsk Rapeseed Research Institute, fertility restorers on sterile cytoplasm were made and estimated for the main valuable traits. The purpose of the research was to develop effective restorers, which are necessary for obtaining of highly productive hybrids on the Polima type cytoplasmic male sterility (CMS). The object of research was 56 varieties of Brassica napus L., which were selected in 2013. It is noted that hybrids F1 were more productive than the parental forms and the standards (the varieties Ratnik and Rif). The restorer line LHR-1 is of practical interest for the development of hybrids on the Polima CMS.

SEARCH OF FERTILITY REGULATORS FOR POLIMA TYPE CYTOPLASMIC MALE STERILITY OF SPRING RAPE (Brassica napus L.) AND ASSESSMENT OF THEIR QUALITY

One of the prior directions in modern rapeseed breeding is making heterotic hybrids. Seed production of most hybrids F1 used in production are often carried out by two types of cytoplasmic male sterility, there are Polima and Ogura. At the All-Russian Rapeseed Research Institute, fertility restorers on sterile cytoplasm were made and estimated for the main valuable characteristics and biochemical properties. The purpose of the study was to create reducing agents necessary for obtaining highly productive hybrids on the Polima type cytoplasmic male sterility (CMS). The object of research was 56 varieties of Brassica napus L., which were chosen in 2013. The setting of experiments, observations, records and analyzes were carried out using the field plot technique, the methodology of field experiments, the method of statistical data processing. It is noted that hybrids F1 had more productivity than the parental forms and the standard (Ratnik and Rif). Promising reducing agents of cytoplasmic male sterility of the Polima type were identified, which ensured complete restoration of fertility in sterile lines with CMS of the Polima type. The reducing agent LHR-1 is of practical interest for the creation of hybrids on the Polima CMS.

The radish Ogura fertility restorer impedes translation elongation along its cognate CMS-causing mRNA

The control of mRNA translation has been increasingly recognized as a key regulatory step for gene control but clear examples in eukaryotes are still scarce. Nucleo-cytoplasmic male sterilities (CMS) represent ideal genetic models to dissect genetic interactions between the mitochondria and the nucleus in plants. This trait is determined by specific mitochondrial genes and is associated with a pollen sterility phenotype that can be suppressed by nuclear genes known as restorer-of-fertility (Rf) genes. In the study, we focused on the Ogura CMS system in rapeseed and showed that the suppression to male sterility by the PPR-B fertility restorer (also called Rfo) occurs through a specific inhibition of the translation of the mitochondria-encoded CMS-causing mRNA orf138. We also demonstrate that PPR-B binds within the coding sequence of orf138 and acts as a ribosome blocker to specifically impede translation elongation along the orf138 mRNA. Rfo is the first recognized fertility restorer shown to act this way. These observations will certainly facilitate the development of synthetic fertility restorers for CMS systems in which efficient natural Rfs are lacking.

Exploring Natural Allelic Variations of the β-Triketone Herbicide Resistance Gene HIS1 for Application in indica Rice and Particularly in Two-Line Hybrid Rice

Background Benzobicyclon (BBC) is a β-triketone herbicide (bTH) used in rice paddy fields. It has the advantages of high efficiency, low toxicity, high crop safety, and good environmental compatibility, and shows efficacy against paddy weeds resistant to other types of herbicides. However, as some important indica rice varieties are susceptible to BBC, BBC is currently only registered and applied in japonica rice cultivation areas. Results By analyzing haplotypes of the bTHs broad-spectrum resistance gene HIS1 and phenotypes for BBC in 493 major indica rice accessions in China, we identified a novel non-functional allelic variant of HIS1 in addition to the previously reported 28-bp deletion. Through detection with markers specific to the two non-functional mutations, it was clear that 25.4% of indica conventional varieties, 59.9% of fertility restorers, and 15.9% of sterile lines were susceptible to BBC. In addition, due to natural allelic variations of the HIS1 gene in the sterile and restorer lines, some two-line hybrid sterile lines were sensitive to bTHs, and the corresponding restorers were resistant. We showed the potential effectiveness of using bTHs to address the issue of two-line hybrid rice seed purity stemming from the self-crossing of sterile lines during hybrid rice seed production. Finally, allelic variations of the HIS1 gene may also play an important role in the mechanized seed production of hybrid rice. Conclusions Our findings offer guidance for the application of BBC in indica rice areas and provide a non-transgenic approach to address the seed purity issue of two-line hybrid rice.

Exploring natural allelic variations of the β-triketone herbicide resistance gene HIS1 for application in indica rice and particularly in two-line hybrid rice

Background: Benzobicyclon (BBC) is a β-triketone herbicide (bTH) used in rice paddy fields. It has the advantages of high efficiency, low toxicity, high crop safety, and good environmental compatibility, and shows efficacy against paddy weeds resistant to other types of herbicides. However, as some important indica rice varieties are susceptible to BBC, BBC is currently only registered and applied in japonica rice cultivation areas.Results: By analyzing haplotypes of the bTHs broad-spectrum resistance gene HIS1 and phenotypes for BBC in 493 major indica rice accessions in China, we identified a novel non-functional allelic variant of HIS1 in addition to the previously reported 28-bp deletion. Through detection with markers specific to the two non-functional mutations, it was clear that 25.4% of indica conventional varieties, 59.9% of fertility restorers, and 15.9% of sterile lines were susceptible to BBC. In addition, due to natural allelic variations of the HIS1 gene in the sterile and restorer lines, some two-line hybrid sterile lines were sensitive to bTHs, and the corresponding restorers were resistant. We showed the potential effectiveness of using bTHs to address the issue of two-line hybrid rice seed purity stemming from the self-crossing of sterile lines during hybrid rice seed production. Finally, allelic variations of the HIS1 gene may also play an important role in the mechanized seed production of hybrid rice.Conclusions: Our findings offer guidance for the application of BBC in indica rice areas and provide a non-transgenic approach to address the seed purity issue of two-line hybrid rice.