Exploitation of hybrid vigour at commercial level through development of hybrid wheat is considered as one of the promising approach for increasing wheat productivity. Work on hybrid wheat started world over in 1962 and utilization of heterosis through hybrid breeding has given very high dividends in many field crops including horticulture.

The discovery of an effective cytoplasmic male sterility and pollen fertility restoration systems in wheat opened up new avenues for commercial hybrid seed production. Kihara (1951) pointed out the possibility of using male sterility by transferring the nucleus of common wheat into the cytoplasm of Aegilops caudata. Among the cytoplasms which in an interaction with Triticum nucleus bring about sterility, T. timopheevii seems to be the most suitable one for commercial production of hybrid seed. Wheat is monoecious and therefore, a line designated as female must not be allowed to produce pollen capable of fertilization while acting as a parent. As wheat flowers are hermaphrodite, the labour requirement for mechanical emasculation is very high. Methods for procuring male sterility in plants may be genetically controlled including cytoplasmic male sterili­ty, nuclear male sterility and self-incompatibility or through chemical hybridization agents or hand emasculation. Three methods for obtaining male sterility have been proposed for wheat based on utilization of genes, i.e., nuclear genes, cytoplasmic and nuclear genes interaction (CMS), chemicals (CHA) or environments (TGMS, PGMS, etc.) but CMS and CHA approaches are widely used in wheat as pollination control system. There are now more than 70 different male sterile cytoplasm systems reported in wheat. 

Genetic male sterility

A number of studies have been conducted for utilization of cytoplasmic male sterility system for hybrid production in wheat but some practical difficulties were proved in hybrid wheat breeding programmes using T-type cytoplasm induced male sterility. Firstly, the level of hybrid vigour was affected by the alien T. timopheevii Zhuk. cytoplasm; and  secondly, the restoration of male sterility in the F1 was genetically complex, incomplete and affected by genetic background. A new pollination control system for producing hybrid wheat seeds by the interaction between a genetic male sterility (GMS) gene and an added alien chromosome from a related species was proposed. Number of genes have been identified that prevent the production of pollen capable of fertilisation. It is possible for male sterility to be achieved without a significant reduction of female fertility. Since most normal wheat varieties will act as restorers of fertility, necessary for the Fl plant, it is only necessary to breed special female parents. Genes controlling nuclear male sterility fall into two categories: those showing dominance or near dominance for sterility and those which are recessive. Fertility in plants is usually encoded by dominant genes. Mutations of a single gene are the probable cause of the majority of examples of Nuclear / genic male sterility in plants. These may cause an alteration of a gene to the recessive state leading to loss of function and sterility. Except in special circumstances, reces­sive genes for male sterility are highly desirable because they usually facilitate identification of homozygous sterile progenies for Fl production. There have been many reports of genetic male sterilty (GMS) of wheat in the literature but only five GMS loci, two recessive (ms1 and ms5) and three dominant (Ms2, Ms3 and Ms4) have been located to wheat chromosomes so far. ms1 and ms5, the recessive genes located to chromosome arm 4BS and 3AL, respectively. The locus ms1 has six designated mutant alleles (ms1a, ms1b, ms1c, ms1d, ms1e and ms1f) whereas locus ms5 has one mutant allele. The six ms1 mutants were named Pugsley’s-ms1a, Probus-ms1b, Cornerstone-ms1c, FS2-ms1d, FS3-ms1e and FS24-ms1f. Two of these mutants were produced by ionizing radiation (Cornerstone and Probus) and are presumed to result from terminal deletions of chromosome arm 4BS.  However Pugsley’s mutant was a spontaneous mutant identified in an F3 progeny of a cross Kenya Farmer/Bearded Javelin 48 in Australia and likely to have an intact 4BS arm. The other four mutants, FS2, FS3, FS20 and FS24, were chemically (EMS) induced and are therefore, likely have intact 4BS arm.  Three dominant male sterile genes Ms2, Ms3 and Ms4 were located to chromosome arm 4DS, 5AS and 4DS, respectively.  

The obstacle to use of NMS systems is maintenance of the male sterile parent. Normally this is achieved within a heterozygous population. Te problem is for wheat because low multiplication rates make hand roguing of fertile segregants uneconomic and unsatisfactory. Maintenance requirements have, therefore, been the main consideration in the systems for NMS production in wheat. The XYZ system and VE system were more commonly used systems conferring genetic male sterility in wheat. Some other genes conferring male sterility were S738, UC9109-9, ‘Yuma’ x ‘Capeiti’ and the LZ mutant.  Despite full investigations, GMS had not been utilised commer­cially for cereal crops. Apart from use in China, there is little to suggest that GMS has yet reached commercial use. The basic requirement for the GMS system is the existence of genes encoding male sterility. The drawback with most systems is large scale production of uniformly sterile seed. However, it offers certain potential benefits including the absence of the need for special restorer varieties and reasonable freedom from the damaging side-effects apparent in many forms of CMS. Most conventional varieties of wheat may be used for this purpose, making it possible to select male hybrid parents from a wide range of varieties. In addition, GMS may also be used in recurrent selection schemes. The disadvantage of GMS wheat was the maintenance the male sterile line. In general, this requires production of a population heterozygous to at the locus controlling fertility. The fertile segregants must then be removed from the hybrid seed production field prior to flowering.

Cytoplasmic genetic male sterility

This system is commonly known as CMS system and involves three types of parental lines namely, the male sterile line (A-line), the maintainer line (B-line) and the fertility restorer line (R-line). Hybrids are the resultant of A x R crosses. Cytoplasmic genetic male sterility is visualized as an essential genetic tool to F1 hybrids in self pollinated crops. Kihara reported the cytoplasmically induced male sterility in wheat for the first time in 1951 by substituting common wheat genome into Aegilops caudata cytoplasm. The possibility of hybrid wheat became apparent after discovery of another effective source of male sterility in T. timopheevi / Bison cross by Wilson and Ross in 1962.  A number of species from genera Triticum and Aegilops were reported as source of male sterility but Triticum dicoccoides var. spontaneovillosum, Ae. aucherii, T. zhukovskii, Ae. Ventricosa, Ae.. kotschyi, Ae. caudata and Ae. ovata were considered as the good sources of cytoplasmic genetic male sterility. Triticum timopheevi was observed to be major source of male sterility in wheat by several workers and is now most widely used cytoplasm in hybrid wheat production. T. timopheevii and its derrivatives were also observed as potential source of fertility restoration system in wheat. Other sources of fertility restoration in wheat were wheat varieties Primpei, Lal Bahadur, Ridley, HD1944 and related hexaploid species T. dicoccoides var. spontaneovillosum and T.spelta var. duhamelianum.

The genetic control of this male sterility was explored by several workers and different reports were made in this regard. Two incomplete dominant genes with epistatic action were observed to control fertility-sterility system in wheat.  A gene ms2 for male sterility was also observed on short arm of chromosome 4D. The male sterility was also reported to be controlled by double recessive nuclear type action and involvement of upto 2 genes, designated as fms1 and fms2 for male sterility was advocated. Single recessive gene was also found responsible for male sterility in wheat. Fertility restoration in wheat has been reported to be under the control of monogenic, digenic as well as polygenic control. The complexity of the genetics of restoration is evident from the fact that restorer genes are carried on many chromosomes i.e., 1, 2, 5, 6 and 7 in wheat. Three genes, located on chromosome 1A, 6B and 7D of Triticum-timopheevii were found responsible for complete fertility restoration. However, all the three (monogenic, digenic and polygenic) mode of inheritance were reported in wheat for fertility restoration. Chromosome 1B of Chinese Spring was also identified as carrier of fertility restoration gene. Multiple recessive genes with minor effects were found responsible for control of fertility restoration in wheat. Interspecific cytoplasmic male sterility and male fertility restoration systems in hexaploid wheat are conditioned by interaction involving dosage of Rf genes (restoration of fertility) and Fi genes (Fertility inhibiting) in polyploidy nucleus and the cytoplasmic genes of the related alloplasmic species. The plant vigour restoring genes Vi also play important role. More than 10 different Rf genes from T. timopheevii, Secale, spelt wheat, emmer and wild emmer, etc. relative species have been discovered and used in breeding R-lines. 

Different sources of male sterility system carry their own negative effects on growth and productivity of wheat. The cytoplasm from Triticum timopheevii has been found to exhibit tendency towards pistiloidy and meiotic instability, reduced dry matter weight, kernel shriveling, low seed germinability, higher anther extrusion, increased tiller production and ear length, decreased grain weight/ear and reduced grain filling, ear sprouting, etc. by several workers. The Aegilopes cytoplasms were also found to express delayed heading, reduced plant height, longer ears, more productive tillers/plants and shrivelled grains when crossed to maintainer/restorer line owing to late maturity.

The CMS system for producing hybrid seed has been described many times in which the production of a single cross hybrid requires three lines, two to provide a female parent and one for the male. On the female side, after a genotype has been selected for combining ability, the first requirement will be to transfer it to a sterilising cytoplasm. In most species, CMS is incorporated into a line of the female parent by backcrossing using the line with the male sterile cytoplasm as the recurrent parent and the normal, male fertile line, as the donor parent. The CMS line is, by convention often designated the ‘A' line. Although it is possible for CMS lines, once developed, to remain true to type, problems are sometimes experienced in the maintenance of sterility.  A line carrying the same nuclear genotype in a fertile cytoplasm, usually from T. aestivum, is used for pollinating the A line for maintenance purposes. This line is known as the 'B' line. The preservation of CMS relies on the non-transmission of the cytoplasm by the pollen of the male. The male parent, in addition to being selected for combining ability, must normally carry genes to restore fertility to the F1 hybrid. Restorer genes are incorporated into the male parent by means of a crossing programme, which may also include backcrossing. Restorer genes are given the designation 'Rf' and it is customary to denote the restorer as the 'R' line. In wheat, the B and R lines are both maintained as normal inbred lines, without difficulty, using conven­tional seed production arrangements. The basic system for producing an FI hybrid by means of CMS is through crossing of A line with R line. This system uses the commonly adopted notation whereby the CMS is termed the' A line', its maintainer with the same nuclear genotype is called the 'B line' and the restorer acting as the male parent of the Fl hybrid is the 'R line'.

Chemically induced male sterility

The limitations of cytoplasmic male sterility system, viz., unstable nature, undesirable linkages and need for use of maintainers have prompted to develop simple and more efficient methods to create male sterility by other means like use of chemicals, mutagens for induction of male sterility. Male sterility induced by chemical hybridizing agents is relatively more convenient to use because there is no need to maintain it, does not require any pre-breeding, fast and relatively easy to implement and many and virtually any combination can be explored. Besides, it has disadvantages of very expensive to develop and register, highly crop growth stage-dependent and weather-dependent, often non-complete sterility in female and somewhat phytotoxic effects. Compared with CMS systems, an effective CHA allows the production of large numbers of parental combinations and permits the evaluation of a number of inbreds for combining ability and/or breeding value. This substantially reduces the time required for hybrid development. More than 40 chemicals have been patented as potential CHAs world over out of which etherel and maleic hydrazide are most commonly used. Complete male sterility by spraying of etherel was observed along with reduction in plant height due to reduced internode length, plant development, yield and increased number of fertile tillers. The female sterility and plant damage by maleic hydrazide was also observed in wheat. A comparative study to study the effect of etherel and MH showed that the etherel was more effective towards reduction in seed set than maleic hydrazide. Other chemicals found effective in causing pollen sterility are, KMS-1, WL84811, LY195259 and CH9701. The Shell compound WL84811 and the Monsanto CHA “GENESIS” are well known current generation of CHAs for inducing male sterility. The appropriate stage for higher efficacy of CHA was reported in several reports at spike length of 7-8 mm, early stem extension stage and early boot stage. Further, the late sown crop was found more responsive to CHA than normal sown crop. The hybrid development process via CHA route involves the induction of male sterility in female lines through chemical spray at appropriate stage and the pollination for seed set.  Attributes of an ideal CHA have been suggested by several authors. These attributes for an ideal CHA includes selective induction of only male sterility with no effect on female fertility, production of readily apparent male steriles, no phytotoxic effect on the treated parent, effective on all genotypes of a species in a wide range of environments, has systemic activity or persistence to sterilize early and late tillers or plants in the field population, flexibility in time of application to overcome adverse weather effects and to permit treatment of many hectares, considerable dosage flexibility to permit a safety margin for application, single treatment for achieving sterility, does not adversely affect F1 seed quality, or F1 seedling or plant vigour, effective on several genera, economical to synthesize and practical to apply  and safe and non-toxic to the environment. While there are a rather large number of chemicals that cause male sterility in wheat, there are very few that meet most of the above requirements. Once a chemical is identified as having some CHA activity, much work needs to be done to identify the optimum time and rate of application. Additionally, application rates can vary with environment and genotype and can be modified with the use of chemical surfactants. Chemical mutagenesis has also been used as an alternate approach to overcome undesirable effects of cytoplasmic male sterility system and to induce male sterility. Sodium azide (NaN3) and ethyl methane sulphonate (EMS) were found as potent chemical mutagen for induction of male sterility.

Environment sensitive male sterility (EMS)

Male sterility is also caused by variations in environmental conditions and specific environments produce male sterility. These may be photoperiod- sensitive cytoplasmic male sterility (PCMS), photoperiod sensitive genic male sterility (PGMS), temperature-sensitive genic male sterility (TGMS), photo-thermo-sensitive genic male sterility (PTGMS) and micronutrient deficiency-Induced male sterility. PCMS was caused by an interaction between Aegilops crassa cytoplasm and Triticum aestivum cv. Norin 26 nucleus. Alloplasmic ‘Norin 26’ showed almost complete male sterility under long-day conditions of 15hours or longer, but male fertility under short-day conditions of 14.5 hours or less. The PCMS is maintained and multiplied by self-pollination under short-day conditions (eg. Natural day length in autumn sown conditions) and hybrid seeds can be produced through out crossing of a PCMS line with a pollen parent under long-day conditions (eg. natural day length in spring sown conditions). In contrast to the CMS system, this system requires only a PCMS line and a pollen parent (restorer line). PGMS was observed when wheat plants were subjected to 10h of photoperiod treatment at a stage when stamen primordia were visible on the first floret of the most advanced spike let of the main shoot that led to the transformation of stamens in to ovaries. Ovules were found to develop on the anther lobes to make plants sterile. Thermo-sensitivity (TGMS) in wheat was reported for producing genic male sterility. Certain wheat varieties carry genes, which render male fertility sensitive to environmental conditions such as reduced temperature. In regions having consistent climates, it might be possible to utilize such effects for production of F1 hybrid seed for use in crop production in different environment. Long photoperiods or higher temperatures or both enhance pollen sterility. Increasing latitude toward the north, photoperiod increases that induce male sterility (PTGMS). Deficiencies of copper, boron and some other micronutrients are also reported to induce male sterility in wheat. High phenotypic variation has been reported in sensitivity to the deficiency of these micronutrients. Very sensitive types are completely male sterile under micronutrient deficiency conditions.

It is apparent from above that wheat has wide range of factors causing male sterility systems. However, CMS and CHA systems were widely used in hybrid wheat production. Studies are underway to identify suitable male sterility system that gives complete and stable male sterility with minimum investments. 


Authors:

SK Singh, Dev Mani, Jaydev Kumar and Deepak Kumar

ICAR-Indian Institute of Wheat & Barley Research, Karnal-132001, India

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