Journal of Heredity Advance Access originally published online on April 20, 2005
Journal of Heredity 2005 96(4):404-409; doi:10.1093/jhered/esi054
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Genetic Analysis of the Species Cytoplasm Specific Gene (scsd) Derived from Durum Wheat
From the Department of Plant Sciences, 470G Loftsgard Hall, North Dakota State University, Fargo, ND 58105
Address correspondence to S. F. Kianian at the address above, or e-mail: s.kianian{at}ndsu.edu.
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Abstract |
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The action of species cytoplasm specific (scs) gene(s) can be observed when a durum (Triticum turgidum L.) nucleus is placed in the Aegilops longissimum S. & M. cytoplasm. This alloplasmic combination, (lo) durum, results in nonviable progeny. A scs gene derived from T. timopheevii Zhuk. (scsti) produced compatibility with the (lo) cytoplasm. The resulting hemizygous (lo) scsti– durum line was male sterile and when crossed to normal durum produced a 1:1 ratio of plump, viable (PV) seeds with scsti and shriveled inviable (SIV) seeds without scsti. In a systematic characterization of durum lines an unusual line was identified that when crossed to (lo) scsti– produced all PV seeds. When planted these PV seeds segregated at a 1:1 ratio of normal vigor plants (NVPs) and low vigor plants (LVPs). The LVP senescence before full maturity. The NVPs were male sterile and when crossed to common durum lines resulted in all plump seeds that again segregated at a 1:1 ratio of NVPs to LVPs. The crosses of these NVPs to common durum lines resulted in a 1:1 ratio of PV to SIV seeds. This study was extended to 317 individuals segregating for scsti and the new locus, derived from durum wheat (scsd), establishing the allelic relationship of these two genes.
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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Durum (Triticum turgidum L. 2n = 28; 14''; AABB) and common wheat (T. aestivum L. 2n = 42; 21''; AABBDD) differ in regard to compatibility with the D- and M-genome Aegilops species (Franckowiak et al. 1976). For example, common wheat is compatible and durum wheat is incompatible with the cytoplasm of Aegilops longissimum S. & M. (2n = 14; 7''; SS hereafter referred to as lo) or A. uniaristata Vis. (2n = 14; 7''; MM hereafter referred to as un) (Maan 1992a, 1992b). Common wheat has certain species cytoplasm specific (scs) gene(s) that are absent in the nuclear genome of durum wheat.
Cytoplasmic genes are maternally inherited. To maintain a cytoplasm, the backcross method can be used to substitute the nuclear genome of one species into the cytoplasm of another (Fukasawa 1953; Kihara 1951; Maan et al. 1999). Alien cytoplasms and chromosomes have been introduced into wheat by cytologically monitoring the chromosomal constitution of backcross progenies in interspecific hybrids (Maan et al. 1999). The cytoplasmic male sterility (CMS) trait is of particular interest in alloplasmic wheat (wheat with an alien cytoplasm) for the production of commercial hybrids (Anderson and Maan 1995; Lucken 1987; Maan and Kianian 2001a). The commercial production of hybrid wheat creates an opportunity for wheat improvement, including advantageous characteristics such as hybrid uniformity, gene pyramiding within a hybrid, and hybrid vigor. However, nuclear-cytoplasmic (NC) incompatibilities have hindered researchers from making many possible hybrid gene transfers in wheat.
There is evidence of NC compatibility and incompatibility reactions in other plant species as well, most often cytoplasmic male sterility (Hanson 1991). Cytoplasmic male sterility has been reported in over 150 plant species and has been consistently found to be associated with the expression of a novel mitochondrial peptide (Leon et al. 1998). An extra peptide is cotranscribed with this novel mitochondrial peptide in Brassica, maize (Zea mays L.), and common bean (Phaseoulus vulgaris L.) (Kennel et al. 1987; Singh and Brown 1991). Examples of these mitochondrial genes in maize are a detoxification protein, encoded by Rf2, which corrects the CMS and nonchromosomal stripe (ncs) mutation phenotypes (Cui et al. 1996; Newton 1995). The ncs mutation phenotypes, which range from infertility or leaf striping to loss of mitochondrial gene function or severe growth impairment, also arise due to NC genetic combinations (Newton 1995). A similar interaction occurs in common bean. The novel polypeptide pvs-orf239 accumulates only in the reproductive tissue of common bean (Wang et al. 1993). However, the mitochondrial genome shifts the pvs-orf239 to substoichiometric levels within the genome if the dominant nuclear factor Fr is added. This interaction results in pollen fertility (Janska and Mackenzie 1993). A final example of these genes is a dominant nuclear gene CHM in Arabidopsis that causes mitochondrial genomic rearrangements (Leon et al. 1998).
Two main obstacles that hinder alien gene transfer in wheat are hybrid sterility and chromosome asynapsis (Maan et al. 1999). Most often genes affecting NC interaction are directly or indirectly involved (Maan 1975). In general, durum is a more sensitive indicator of alloplasmic compatibilities than hexaploid wheat. In certain alloplasmic situations where T. aestivum is self-fertile, T. turgidum may only be partially fertile or even completely sterile (Maan 1992b).
Restoration of fertility (Rf) genes have traditionally been used to restore fertility in sterile hybrids; however, these genes do not restore fertility in all situations (Maan 1992b). One circumstance in particular involves incompatibility between A. longissimum cytoplasm and the T. turgidum nucleus. A two-gene system has been found to restore fertility in this and other cases where Rf genes do not restore fertility. The genes involved in this system are the species cytoplasm specific (scs) gene and the vitality (Vi) gene described by Maan (1992c).
Compatibility with the A. longissimum cytoplasm (lo) is evaluated by the recovery of plump viable (PV) and shriveled inviable (SIV) seeds. The presence of PV seeds indicates compatibility; this is the case with a T. aestivum nucleus in an A. longissimum cytoplasm. The presence of SIV seeds characterizes incompatibility, as found in (lo) durum (Maan 1992a). The scsti gene was introgressed into the durum background from the nuclear genome of T. timopheevii Zhuk. (Maan 1983). This gene improved nuclear cytoplasmic and embryo–endosperm compatibility between alloplasmic T. turgidum and A. longissimum. Expression of the scsti gene is demonstrated as seed viability and partial compatibility between the durum nucleus and (lo) cytoplasm producing normal vigor but male-sterile plants (Maan 1992a). The scs genes from respective cytoplasm donors nearly mitigate cytoplasmic effects, but not completely. This is where the Vi gene becomes fundamental in the compatibility system. The Vi gene restores NC compatibility, embryo–endosperm compatibility, seed viability, and fertility to (lo) durum plants containing scs (Maan 1992c). However, the Vi gene alone does not restore plant vigor. It is hypothesized that the Vi gene resulted from a spontaneous mutation of an Rf gene located on the short arm of chromosome 1B (Maan 1992b).
Maan (1994) has investigated the actions of the scsti and Vi genes and the interactions between these two loci. In the absence of Vi, alloplasmic plants containing the scsti gene have normal plant vigor, are male-sterile, and produce a 1:1 ratio of PV to SIV seed with and without scsti, respectively, when crossed to durum containing no scs gene. Those plants containing Vi but not scsti produce fertile plants with reduced vigor. Plants containing both the scsti and Vi genes produce fertile plants with normal growth and vigor. In the absence of both genes the T. turgidum nucleus is incompatible with the A. longissimum cytoplasm. A similar situation exists with A. uniaristata cytoplasm (Maan 1975).
Several scs genes have been identified that produce compatibility between durum wheat and the (lo) or (un) cytoplasm. These include scsae on the long arm of chromosome 1D (1DL) from common wheat, scsun on a telocentric chromosome from A. uniaristata that is homologous to group 4, scsti from T. timopheevii located on the long arm of chromosome 1A (1AL; Anderson and Maan 1995), and two genes, one located on a 2S.2SL translocation chromosome having the short arm of chromosome 2 from A. speltoides and a second on a 4S.4AL translocation chromosome having 4S from A. longissimum (Maan and Kianian, 2001b).
In a systematic characterization of durum lines, an unusual line was identified that produced all PV seeds when crossed to (lo) scsti–. When these seeds were planted they segregated at a 1:1 ratio of normal vigor plants (NVPs) and low vigor plants (LVPs). The objectives of this study were to ascertain the genetic and allelic nature of NVP:LVP and PV:SIV seed set in segregating populations.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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A hemizygous (lo) durum line having a scsti gene on chromosome 1A was available from Maan et al. (1999). This line is male sterile and maintained by crossing to normal durum [(d) – –]. Each successive cross produces PV seed having scsti and SIV seed without scsti.
The 28-chromosome unusual euplasmic durum line hypothesized to contain a new scs locus, designated scsd, was selected from the progeny of a durum line that produced all PV seed when crossed to (lo) scsti–. This line has the designation of Line-1 or 00A550-3 but is more commonly referred to as scsd line. Production of all PV seeds indicated the presence of an scs-like gene in the durum parent (scsd). Planting the PV seeds from this cross resulted in male-sterile plants with a 1:1 ratio of NVPs to LVPs.
Allelism between the scsti and scsd genes was assessed in the segregating generations of populations and lines derived from the cross of (lo) scsti– to (d) scsd scsd. A total of 317 individuals were tested over three greenhouse seasons with two populations in fall 2000, three populations in spring 2001, and two populations in fall 2001. Greenhouse conditions were controlled to optimize plant growth for durum wheat and reduce environmental stress. Each plant was evaluated for vigor and classified as either NVP or LVP. All NVPs then had at least two spikes crossed to normal durum [(d) – –]. These crosses were assessed for formation of PV and SIV seed. Resulting ratios of NVPs to LVPs and PV to SIV seed sets were tested using a chi-squared test (Steel et al. 1997) and a confidence level of 95% or greater.
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Results and Discussion |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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Evaluation of first, second, and third segregating generations of the (lo) scsti– x (d) scsd scsd population (Figure 1) allowed analysis of plant genotypes as well as investigation of the possibility of an allelic relationship between the scs gene derived from T. timopheevii (scsti) and the gene discovered from durum wheat (scsd). Genotypes of individuals in these populations were determined based on seed set ratios of plump to shriveled seed and plant vigor.
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Alloplasmic (lo) scsti– durum plants were crossed to euplasmic (true cytoplasm) durum lines having a scsd gene pair [(d) scsd scsd] (Figure 1). Resulting progeny in this first segregating generation were evaluated for plant vigor (Figure 2). Normal plants from this first segregating generation were crossed to durum with no scs gene [(d) – –], creating the second segregating generation (Figure 1). The resulting seed was evaluated for PV to SIV seed ratios and plump seeds were sown and evaluated for plant vigor (Figure 3). Normal vigor plants were crossed again to (d) – – (Figure 1) to create the third segregating generation. The plants were then evaluated for PV to SIV seed set ratios.
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As is evident from the initial experiments, the scsd gene alone restores seed plumpness and therefore partial compatibility in the (lo) cytoplasm, just as the scsti gene does. Unlike the scsti gene however, the scsd gene alone is unable to restore plant vigor in the alloplasmic condition. Chi-square analysis indicated an allelic interaction based on a 1:1 ratio of normal to weak vigor plants in the first and second segregating generations (Table 1). Ratios fit a 1:1 ratio of normal to weak vigor plants with a 95% level of confidence.
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The NVPs from the first segregating generation were crossed to a durum line known not to contain any scs gene [(d) – –; Figure 1A]. Again, the presence of an allelic scs gene pair was indicated by the development of all plump seed (Figure 1A; Table 2). Taken together, the results of this and previous generations indicate that scsti and scsd are alleles of the same locus. Even in the case of close linkage, a few SIV seeds would be expected. Overall, 918 seeds from 7 plants were produced for the above generation, and no SIV seeds were identified.
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When planted these seeds produced a second segregating generation of NVP and LVP in a 1:1 ratio (Figure 1, Table 1). As previous, NVP were crossed to (d) – – and seed formation was evaluated. Assuming the genes are allelic, these crosses were expected to result in a 1:1 ratio of PV to SIV seed. Slight deviation from this ratio, at P = .05 (
) was found in five of the seven populations evaluated; population 2 (
2 = 8.32), population 4 (2 = 8.30), population 5 (2 = 14.01), population 6 (2 = 14.57), and population 7 (2 = 10.27) (Table 2). However, seed set ratios did not fit any other expected (i.e., nonallelic situation) or unexpected ratios (Table 2). It is possible that these deviations could be of a random nature or conceivably the nature of the scs gene itself. For the plant vigor to segregate in a 1:1 ratio and produce a 1 PV:1 SIV seed set ratio, the scsti gene must be allelic to the scsd gene. The original female (lo) parent was known to be hemizygous for the scsti gene; therefore the original durum parent must have been homozygous for the scsd allele. This is expected for durum is a self-pollinating species. Through this schematic (Figure 1A) it can be deduced that the first generation of individuals segregated NVP that were compound heterozygotes with both the scsti and scsd alleles; and LVP that contained only the scsd allele. When these NVP were crossed to (d) – –, that result was all PV due to the compatibility restoration reaction induced by the presence of either the scsti or scsd gene. This cross resulted in NVP hemizygous for the scsti gene and LVP hemizygous for the scsd gene. A nonallelic reaction (Figure 1B) would have resulted in a 3:1 ratio of PV to SIV seed in the first segregating generation, not all plump as observed. When planted, the plump seeds from a nonallelic interaction also would have resulted in three genotypes displaying a 2:1 ratio of NVPs to LVPs. Overall, four distinct genotypes in a ratio of 1:1:1:1 would have resulted in the second segregating generation [(lo) scsti–; scsd–, (lo) scsti–; – –, (lo) – –; scsd–, and (lo) – –; – –]. Normal vigor plants with the genotype (lo) scsti– ; scsd– would produce a 3:1 ratio of PV:SIV seed, while the remaining (lo) scsti– ; – – plants would produce a 1:1 ratio of PV:SIV seed. The results obtained do not match the expected results for a nonallelic interaction. Therefore we can conclusively confirm that scsti and scsd are alleles of the same locus.
In the second segregating generation, at least two spikes from each male-sterile, normal vigor (lo) scsti– plant were crossed. In a few cases, spikes on the same plant deviated from the expected ratio. These deviant individuals were observed in all three greenhouse seasons. In all, 26 of the 317 individual crossed spikes developed deviating seed set ratios. Of the 26 individual spikes found to deviate from the expected 1:1 ratio of PV to SIV seed, 13 of the deviants were from individual spikes where two spikes were crossed per plant (Table 3). Five individuals were found to deviate from the 1:1 ratio of PV to SIV seed set in both spikes, when two spikes were crossed per plant. This is representative of 10 individual spikes of a deviating nature. Three individual spikes, one from each plant, were found to deviate where three spikes were crossed on a plant. The number of plants that produced more PV seed were also compared to plants that produced more SIV seed. These results demonstrate how several different phenotypes are being expressed in an individual plant/spike. Tendency is toward formation of a functional scs (scsti or scsd) resulting in PV seed, as indicated by 22 of the 26 individuals that produced more PV seed than SIV seed, compared to only four individuals that produced more SIV than PV seed (Table 3).
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Deviation from the expected 1:1 ratio of PV to SIV seed and deviations in one spike out of two or three spikes crossed could be an indication of genetic instability with regard to scsti. Another variation noted in the seed set was the observation of a "blistering" phenotype. This phenotype could be described as partially plump and partially shriveled or blistered seed. Environmental influences can be excluded as a cause of inducing this phenotype, because not all the seed from a single cross exhibited blistering, nor did the seed from adjacent plants. Also, the same blistering phenotype was observed by Simons et al. (2002) in a population developed for the fine structure mapping of the scsti gene. These observations indicate the influence of other factors on the expression of the scs genes.
The blistering phenotype was initially observed in maize with regard to the maize brittle-1 mutable (bt-1m) gene (Sullivan et al. 1991). The normal bt-1 gene produces seed normal in appearance, whereas mutated bt-1 produces a kernel shrunken in appearance. The bt-1m gene produces a blistering phenotype. Through genetic and molecular analyses, it was ascertained that the bt-1m was the result of an insertion of a defective suppressor-mutator (dSpm) transposable element (Sullivan et al. 1991). The similarity between the blistering phenotype in maize and the blistering phenotype associated with the scsti gene may suggest the presence of active transposable element(s) causing the development of different seed phenotypes. Observation of a blistering phenotype may indicate possible sectoring of tissue in a single seed consistent with the phenotype observed when active transposable elements are detected.
Additional evidence is provided from the research by Simons et al. (2002). In a population of 110 plants, 11 individuals were found exhibiting double crossovers within a 3-cM region. This was much higher than expected. To explain this deviation from the expected observation of two double-crossover individuals, Simons et al. (2002) concluded that active transposable elements in the scs locus were responsible for changes in its allelic state.
Further research into the scsd gene and the discrepancies produced in seed set ratios is possible by planting remaining seed of those lines that produced deviating ratios. Remnant seed from deviating individuals has been planted and will be evaluated to check if deviating seed set ratios are again observed. The phenotype(s) exhibited by these individuals could provide evidence to either support or reject the possibility of an active transposable element through the observance of subsequent deviating ratios from those plants where deviating ratios were initially observed.
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Acknowledgments |
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We thank Justin B. Hegstad, Kay M. Carlson, and the rest of the Wheat Germplasm Enhancement group for their assistance in making this article possible. We also thank Drs. Christoffers and Cai for their suggestions in improving this manuscript, and Dr. R. L. Phillips for helpful discussion and suggestions on bt-1m blistering phenotype of maize. This material is based on the work supported by the USDA-IFAFS grant no. 2001-52100-11293, NSF-PGRP contract agreement no. DBI-9975989, and the North Dakota Wheat Commission to S.F.K.
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Footnotes |
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Corresponding Editor: J. Perry Gustafson
Received May 14, 2004
Accepted January 31, 2005
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References |
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