Genetics of thermosensitive genic male sterility in rice

Summary Inheritance of thermosensitive genic male sterility (TGMS) in Norin PL12 and IR32364TGMS and their allelic relationship were studied from F₁, F₂ testcross (TC) and F₃ generations of the crosses made with the two mutants and several fertile tester parents. F₂, TC and F₃ segregation behavior for pollen and spikelet fertility indicated that the TGMS trait in the two mutants was controlled by a single recessive gene. Allelic relationship studies indicated that TGMS genes of the two mutants were different. Since TGMS gene in Norin PL12 has been designated as tms ₂ , the TGMS gene present in IR32364TGMS is tentatively designated as tms ₃ (t) until allelic test is done with another TGMS gene (tms ₁ ) reported from China in a line 5460S seeds of which were not available.     

Development of TGMS lines for developing two-line rice hybrids for the tropics

A breeding program was initiated at the International Rice Research Institute (IRRI) in 1990 to develop thermosensitive genic male sterile (TGMS) rice lines for developing two-line rice hybrids for the tropics. The TGMS trait was transferred from a temperate japonica TGMS mutant, Norin PL 12, to indica and tropical japonica rice varieties using the pedigree selection procedure. Six new TGMS rice lines adapted to tropical conditions were developed which showed complete pollen and spikelet sterility when maximum temperature was higher than 30 ^°C 1–2 week after panicle initiation. However, up to 85.5% spikelet fertility was observed when these lines were exposed to 26–29 ^°C during the critical stage. Using two of these TGMS lines, some heterotic rice hybrids showing 1–1.6 t/ha higher grain yield than the inbred check varieties were identified in unreplicated observational yield trial conducted at IRRI. Two of the six two-line hybrids yielded significantly higher than the check variety in a replicated preliminary yield trial. This revised version was published online in July 2006 with corrections to the Cover Date.     

Expression of thermosensitive genic male sterility in rice under varying temperature situations

Response of thermosensitive genic male sterility in rice to varying temperature situations was studied by using four TGMS lines. In three sets of maximum, minimum and their combined temperatures, it was observed that maximum temperature played a predominant role in influencing sterility/fertility of TGMS lines under the combined regimes. Expression of a TGMS gene was found to be influenced by the genetic background of the recipient lines. Exposure for more than 8 hours of 32 ^°C was found necessary to induce complete male sterility in indica TGMS lines: IR68945-4-33-4-14 and IR68949-11-5-31 while, more than 4 hours of such exposure was enough to induce sterility in case of their japonica donor Norin PL 12. Sudden interruption with 27 ^°C even for 2 hours under the sterility inducing regime of 32/24 ^°C could induce partial fertility in the line IR68945-4-33-4-14. However, the line ID 24 remained completely sterile even with 10 hours of interruption with 27 ^°C. An interruption with lower temperature of 27 ^°C for 4 hours for even one day induced partial fertility in IR68945-4-33-4-14. The period of four to eight days after panicle initiation was the stage most sensitive to temperature variations. Hybrid rice breeders need to develop numerous genetically diverse TGMS lines, which possess critical sterility inducing temperature of 28 ^°C and are not affected by sudden interruptions with a lower temperature for few hours daily for a couple of days. This revised version was published online in August 2006 with corrections to the Cover Date.     

Pleiotropic effect of the dwarfing gene d18-k on cool tolerance at booting stage under the genetic background of an extremely cool-tolerant line Norin-PL8 in rice

Norin‐PL8 (‘PL8’) is an extremely cool‐tolerant line of rice in Japan that contains genes for cool tolerance originating from a javanica landrace. It was investigated to see whether the dwarfing gene d18‐k (kotaketamanishiki dwarf) exerts its pleiotropic effect on enhancing the cool tolerance at the booting stage in the genetic background of PL8. The d18‐k isogenic line of the recurrent parent PL8 (D8), PL8, and two commercial cultivars ‘Hayayuki’ and ‘Kirara 397’ were used. For each line/cultivar, the 12°C‐5‐day treatment was conducted at various times during the booting stage. In addition to spikelet fertility, the ratio (%) of the fertilized‐spikelet number of each treated panicle to the varietal mean of fertilized‐spikelet number per panicle in the control (FS‐T/C) was adopted to estimate cool temperature damage. For FS‐T/C, the lines‐cultivars ranked in the order of D8 > PL8 > ‘Hayayuki’ > ‘Kirara 397’, reflecting their cool tolerances. D8 exceeded PL8 in both pollen grain number per anther in the control and as an indicator of pollen fertility after the treatment, as a result of the effects of d18‐k. Consequently, d18‐k can be used to develop super‐highly cool‐tolerant cultivars for cool‐weather areas.     

Introgression of Xa4, Xa7 and Xa21 for resistance to bacterial blight in thermosensitive genetic male sterile rice (Oryza sativa L.) for the development of two-line hybrids

Bacterial blight (BB) of rice caused by X. oryzae pv. oryzae is a major production constraint in commercial hybrid rice production in the Philippines because most of the parental lines used in hybrid production do not carry resistance genes against the pathogen. In this study, three bacterial blight resistance genes, Xa4, Xa7 and Xa21, were introgressed to a temperature-sensitive genetic male sterile (TGMS1) line. A three-way cross of AR32-19-3-3/TGMS1//IRBB4/7 (PR36944) was made to produce 1,364 F₂ plants carrying various combinations of Xa4, Xa7 and Xa21. Individual plants were characterized for reaction to bacterial blight PXO61 (race 1), PXO86 (race 2), PXO99 (race 6) and pollen sterility. Of 144 F₂ plants demonstrating resistance against PXO61, PXO86 and PXO99, 22 exhibited highly resistant phenotypes with mean lesion lengths ranging from 0.37–2.97 cm. Analysis of disease reaction identified 20 potential TGMS F₂ plants containing Xa4, Xa7 and Xa21 while 78 plants with Xa4 + Xa7. Phenotypic and polymerase chain reaction (PCR) analyses confirmed PR36944-450, PR36944-473 and PR36944-700 as homozygous for Xa7 and Xa21 and highly resistant to all three Xoo races. Fertility of PR36944-450 and PR36944-700 was restored at permissive temperature in a growth chamber. BB-resistant TGMS lines should facilitate breeding two-line hybrids in the tropics.     

Molecular detection of WA‐CMS restorers from tropical japonica‐derived lines,their evaluation for fertility restoration and adaptation

Hybrid rice based on wild‐abortive cytoplasmic male sterility (WA‐CMS) is important in boosting rice production, which requires diverse parents to harness heterosis. For this, exploiting the diversity of japonica through tropical japonica (TRJ) lines is an excellent route. In this study, 310 TRJ‐based new plant type (NPT) lines were developed and evaluated for Rf3 and Rf4 genes. Gene‐based (DRRM‐Rf3‐5 and DRRM‐Rf3‐10) and functional marker (RMS‐SF21‐5) targeted Rf3 locus, while gene‐linked (RM6100) and functional marker (RMS‐PPR9‐1) targeted the Rf4 locus. The frequency of the restorer allele of Rf3 gene was lower when compared to that of Rf4. Combined phenotypic and molecular screening using gene‐based and functional markers identified 42 lines that carried Rf3 and/or Rf4 genes. All the selected lines produced fertile F₁s when crossed to a WA‐CMS line, “Pusa 6A”, but with varying levels of spikelet fertility. This is the first report of a marker‐cum‐phenotype‐based restorer selection using TRJ‐derived lines. Multilocation evaluation of these lines at three locations indicated better adaptation for grain yield in some of the lines.     

Identification of QTLs for the resistance to rice stripe virus in the <Emphasis Type="Italic">indica</Emphasis> rice variety Dular

The indica variety Dular has a high level of resistance to rice stripe virus (RSV). We performed quantitative trait locus (QTL) analysis for RSV resistance using 226 F₂ clonal lines at the seedling stage derived from a cross between the susceptible japonica variety Balilla and the resistant indica variety Dular with two evaluation criteria, infection rate (IR) and disease rating index (DRI). The experiments were performed in both 2004 and 2005. Based on IR, three putative QTLs were detected and had consistent locations in the 2 years, one QTL was detected in the RM7324–RM3586 interval on chromosome 3. The other two QTLs were linked and located in the RM287–RM209 and RM209–RM21 intervals on the long arm of chromosome 11, and accounted for 87.8–57.8% of the total phenotypic variation in both years. Based on DRI, three putative QTLs were also detected and had consistent locations in both years. One of them was located in the RM1124–SSR20 interval on the short arm of chromosome 11, while the other two linked QTLs had the same chromosomal locations on chromosome 11 as those detected by IR, and accounted for 55.7–42.9% of total phenotypic variation in both years. In comparison to the mapping results from previous studies, one of the two linked QTLs had a chromosomal location that was similar to Stv-b ⁱ , an important RSV resistance gene, while the other appeared to be a newly reported one.     

QTL mapping of male sterility-related traits in a photoperiod and temperature-sensitive genic male sterile wheat line BS366

The photoperiod/temperature-sensitive genic male sterile (P/TGMS) character is important for use of heterosis in hybrid wheat. In the present study, the fertility of 172 doubled haploid (DH) lines derived from the cross between a P/TGMS wheat line BS366 and a restorer wheat line Baiyu149 was investigated under both fertile and sterile environments during three cropping seasons. The phenotype was recorded based on the international seed setting rate. A total of 167 SSR and 1,278 SNP markers were used to construct a linkage map, with a total length of 3,748.94 cM and an average marker interval of 2.59 cM. Three QTLs were identified and designated as QF.bhw-2DS, QF.bhw-4BS and QF.bhw-7Al, explaining 6.9%–12.8%, 19.7%–25.6% and 7.2%–8.8% of the phenotypic variances, respectively. These results lay a good basis for application of male sterility-related molecular markers in improvement of two-line hybrid wheat breeding system.     

Classification of GA response,Rht genes and culm length in Japanese varieties and landraces of wheat

Summary The GA response, Rht genes and culm length of 133 Norin varieties, 6 breeding lines and 16 landraces of Japanese wheat were investigated. Out of 133 Norin varieties tested, 103 were GA-insensitive and 30 GA-responsive. The 6 breeding lines were all GA-insensitive. Out of 16 landraces tested, 10 were GA-insensitive and 6 GA-responsive. Among the 10 GA-insensitive landraces, only Daruma had a Rht1 genotype. The other 9 had a Rht2 genotype. None of the landraces tested carried both Rht1 and Rht2 or Rht3. Out of the 103 GA-insensitive Norin varieties, 22 carried only Rht1, another 79 carried only Rht2, and only Norin 10 and Kokeshikomugi carried both Rht1 and Rht2. No tested variety carried Rht3. Some Norin varieties carrying Rht2 showed tall culms comparable to that of the rht tester line Chinese Spring. These results suggest that these varieties had a nullifier or modifier gene(s) or height promoting genes in the background controlling the height-reducing effect of Rht2. Conversely, six GA-responsive Norin varieties were as short as Akakomugi which carries the GA-responsive Rht genes, Rht8 and Rht9. The also seemed to carry a GA-responsive Rht gene or genes, and moreover all but one may carry gene(s) other than the Akakomugi genes. The origin of Rht1 and Rht2 of Norin 10 was speculated on the GA-response and Rht genotypes of its related varieties and landraces.     

Inheritance and mapping of male sterility restoration gene in upland japonica restorer lines

Several upland Japonica breeding lines, WAB450-11-1-3-P40-HB (Abbreviated as WAB450-11), WAB450-11-1-2-P61-HB (WAB450-13), WAB450-l-B-P-91-HB (WAB450-14), IRAT216, IRAT359, and IRAT104, possessing restoring ability for the Dian 1 type cms (cms-D) line Dianyu 1A were recently identified at Food Crops Research Institute, Yunnan Academy of Agricultural Sciences, P. R. China. In this study, the inheritance of restoring ability in these lines was characterized through the production of backcross populations to the male-sterile and maintainer Dianyu 1 lines. Each of the restorer lines was used to pollinate Dianyu 1A to form a F₁ hybrid which was then backcrossed (1) with Dianyu 1B producing a BC₁F₁ population and (2) to the female parent Dianyu 1A producing a BC₅F₂ population. The lines were also crossed with the japonica restorer line C57, carrying the restorer gene Rf1 that was introgressed from indica, to form F₁ hybrids, these hybrids were then testcrossed with Dianyu 1A to study the allelic relationship of their restorer genes to Rf1. The inheritance in these testcross populations indicated that the complete restoring ability of WAB450-11, WAB450-13, WAB450-14, IRAT216, IRAT359, and the partial restoring ability of IRAT104 were controlled by dominant genes, and the gene in WAB450-13, WAB450-14, and IRAT216 was allelic or identical to Rf1. When 136 SSR markers were used to score 143 BC₁F₁ individuals from Dianyu 1A/WAB450-13//Dianyu 1B, the japonica Rf1 allele was found to be located between RM171 and RM6100 on the long arm of chromosome 10, an interval corresponding to that known for the indica Rf1 allele. The distance between RM171 and Rf1 is 2.8 cM, and that between Rf1 and RM6100 is 4.9 cM. Similar linkage results were obtained from mapping 89 individuals of the corresponding BC₅F₂ population (Dianyu 1A/6/Dianyu 1A/WAB450-13).     

QTLs identification of crude fat content in brown rice and its genetic basis analysis using DH and two backcross populations

Fat content is a concern for the enhancement of rice for eating, cooking, and storage qualities. To clarify its genetic mechanism, a double haploid (DH) population derived from anther hybrid F₁ of Zhenshan 97B (indica) and Wuyujing 2 (japonica) and two backcross F₁ (BCF₁) populations, which came from the DH lines backcrossing to two parents, were used to scan quantitative trait loci (QTLs) and dissect gene effects for the crude fat content (CFC) in brown rice. Fourteen QTLs were resolved, distributing on chromosomes 1, 3, and 5–9. Three loci were detected repeatedly in two populations, DH or BCF₁. Among these loci, a major QTL, qCFC5, flanking markers RM87 and RM334, was located on chromosome 5, which was detected simultaneously among three populations. The main QTLs had a major role in controlling CFC in brown rice and were modified by several mini-effect QTLs and epistatic affection. Wenjun Liu and Jing Zeng are contributed equally to this paper.     

Comparison of two fertility restoration systems against photoperiod-sensitive cytoplasmic male sterility in wheat

A ‘two‐line system’ using photoperiod‐sensitive cytoplasmic male sterility (PCMS) caused by Aegilops crassa cytoplasm under a long‐day photoperiod ( 15 h) has been proposed as a new means of producing hybrid varieties in common wheat. The PCMS line is maintained by self‐pollination under short‐day conditions, and hybrid seeds can be produced through outcrossing of the PCMS line with a pollinator under long‐day conditions. Two kinds of fertility restoration systems against the PCMS are known. One is involved with a set of multiple fertility‐restoring (Rf) genes in the wheat cultivar ‘Norin 61’ located on (at least) chromosomes 4A, 1D, 3D and 5D. The other is controlled by a single dominant major Rf gene, Rfd1, located on the long arm of chromosome 7B in the wheat cultivar ‘Chinese Spring’. To examine the degree of fertility restoration by these two systems, nine PCMS lines were crossed with ‘Norin 61’ and ‘Chinese Spring’ as the restorer lines, and the F₁ hybrids were investigated. The degree of fertility restoration was estimated by comparing the seed set rates in the F₁ hybrids having the Ae. crassa cytoplasm and those with normal cytoplasm. The results revealed that the fertility restoration ability of a set of multiple Rf genes in ‘Norin 61’ was higher than that of the Rfd1 gene in ‘Chinese Spring’.     

Genetic analysis and fine mapping of tms9, a novel thermosensitive genic male‐sterile gene in rice (Oryza sativa L.)

Two‐line hybrid rice as a novel hybrid breeding method has huge potential for yield increasing and utilization of intersubspecific heterosis, and it is of major significance for the food security of rice‐consuming populations. Zhu1S is a thermosensitive genic male‐sterile line of rice with low critical temperature and excellent combining ability, which has been widely exploited as a female parent in Chinese two‐line hybrid rice breeding. Here, genetic mapping in F₂ populations was used to show that its male sterility is inherited as a single recessive gene and that responsible gene (termed tms9) lies on the short arm of chromosome 2. A high‐resolution linkage analysis which was based on the Zhu1S/R173 F₂ population found that the thermosensitive genic male‐sterile gene tms9 of Zhu1S was fine mapped between insertion–deletion (Indel) markers Indel 37 and Indel 57, and the genetic distance from the tms9 to the two markers was 0.12 and 0.31 cM, respectively. The physical distance between the two markers was about 107.2 kb. Sequence annotation databases showed that the two Indel markers (Indel 37 and Indel 57) were located on two BAC clones (B1307A11 and P0027A02). There are sixteen open reading frames (ORF) present in this region. The results of this study are of great significance for further cloning tms9 and molecular marker–assisted selection.     

Identification,distribution and effects on agronomic traits of the semi-dwarfing <Emphasis Type="Italic">Rht</Emphasis> alleles in Bulgarian common wheat cultivars

Bulgarian common wheat cultivars released in the period 1925–2003 were studied using the gibberellic acid (GA) test and microsatellite analysis of the Xgwm261 locus on chromosome 2DS to identify the semi-dwarfing (Rht) genes. The old cultivars, isolated through selection from landraces, carried rare alleles (211- and 215-bp) at Xgwm261 locus, and those developed by hybridisation to foreign cultivars, carried the 165- and 174-bp alleles. Forty-two (55.3%) of 76 modern cultivars were GA-responsive. The 192-bp allele, diagnostic for Rht8, was observed in 64 (84.2%) modern cultivars, of which 37 carry Rht8 alone, and 27 possess a combination of Rht8 and a GA-insensitive allele viz. Rht-B1d (17); Rht-D1b (6) and Rht-B1b (4). The 174-bp allele is present in seven cultivars, only one of which is photoperiod-sensitive, and the rest are day-length insensitive. The 203-bp allele was found in six modern cultivars. Cultivars carrying the Rht8 allele are the most widespread and some of them have been cultivated for a long period. Cultivars with the `Saitama 27' allele (Rht-B1d) are the most productive and are second in distribution in the country. The recently observed trend for increasing the proportion of cultivars with GA-insensitive Rht genes is probably due to their combination with the 192-bp allele of Xgwm261 locus tightly linked to the Ppd-D1, to the break of the link between the 174-bp allele and ppd-D1, and to the introduction of other genes influencing flowering time.     

Identification and application of major quantitative trait loci for panicle length in rice (Oryza sativa) through single‐segment substitution lines

Panicle length (PL), an important yield‐related trait, strongly affects yield components, such as grain number, grain density and rice quality. More than 200 panicle length quantitative trait loci (PL QTLs) are identified, but only a small number are applied in rice breeding. In this study, we performed QTL analysis for PL using 42 single‐segment substitution lines (SSSLs) derived from nine donors in the genetic background of HJX74. Fourteen QTLs and five heterosis QTLs (HQTLs) for PL were recognised. Three QTLs and four HQTLs acted positively, and the other eleven QTLs and one HQTL acted negatively. By scanning the single heterozygous background region of the F₂ population with large‐genetic‐effect SSSLs, we mapped PL loci qPL6‐2 and qPL7‐1 to different locations on chromosomes 6 and 7, respectively, in three consecutive years of independent trials. The genetic effects of these QTLs were further assessed. qPL6‐2 demonstrated the most positive additive effect (QTL), whereas qPL7‐1 achieved the most positive dominant effect (HQTL) for PL. These results indicated that the pyramiding of PL QTLs might increase grain yield and facilitate the application of the beneficial allele in hybrid rice breeding.     

Genetic analysis of fertility restoration against photoperiod-sensitive cytoplasmic male sterility in Triticum aestivum cv. Norin 61

Triticum aestivum cv. Norin 26 with the Aegilops crassa cytoplasm becomes almost completely male sterile when grown under a long-day condition (15 h of light or longer), but is highly male fertile under a short-day condition (14.5h or less). This type of male sterility is called photoperiod-sensitive cytoplasmic male sterility (PCMS). Genetic analyses were made of the fertility-restoring (Rf) genes effective against PCMS that are present in T. aestivum cv. Norin 61. Conventional and monosomic studies indicated that restoration of fertility is controlled by multiple Rf genes located on at least four chromosomes: 4 A, 1D, 3D and 5D. The genetic mechanism of fertility restoration by the genes of‘Norin 61’differs from the mechanisms reported for‘Chinese Spring’and a‘Norin 26’mutant line.     

Distribution of microsatellite alleles linked to Rht8 dwarfing gene in wheat

A wheat microsatellite locus, Xgwm 261, whose 192-bp allele closelylinked to the dwarfing gene Rht8, on chromosome 2D, was used toscreen 71 wheat cultivars from 13 countries to assess the variation at thislocus. Screening of this wheat collection showed that a 165-bp allele anda 174-bp allele were the most frequent. None of the New Zealand cultivarspossessed a 192-bp allele specific to Rht8, while only one cultivarfrom the US produced this important allele. The frequency of a 192-bpallele among these wheat cultivars was 5.63%. The highest allelefrequency was observed for a 174-bp fragment (52.11%) followed by a165-bp fragment (26.76%). The only durum wheat `Cham 1', did notshow any amplification due to the absence of D genome. Four new novelalleles, 180-bp, 198-bp, 200-bp and 204-bp present in the US and NewZealand wheat cultivars are reported.     

Validation of molecular markers linked to fertility restorer gene(s) for WA-CMS lines of rice

The present study was carried out with the objective to validate the molecular markers, which have been previously reported to be linked to fertility restorer (Rf) gene(s) for WA-CMS lines of rice. Two mapping populations involving fertility restorer lines for WA-cytoplasm, viz., (i) an F₂ population derived from the cross IR58025A/KMR3R consisting of 347 plants and (ii) a BC₁F₁ population derived from the cross IR62829A/IR10198R//IR62829A consisting of 130 plants were analyzed. Nine SSR and three CAPS markers reported to be linked to Rf genes along with two previously unreported SSR markers were analyzed in the mapping populations. In both the populations studied, the trait of fertility restoration was observed to be under digenic control. Eight SSR markers (RM6100, RM228, RM171, RM216, RM474, RM311, MRG4456 and pRf1&2) showed polymorphism between the parents of the F₂ population, while the SSR markers RM6100 and RM474 showed polymorphism between the parents of both the F₂ and BC₁F₁ populations. Only one CAPS marker, RG146FL/RL was polymorphic between the parents of the BC₁F₁ population. RM6100 was observed to be closely segregating with fertility restoration in both the mapping populations and was located at a distance of ~1.2 cM. The largest phenotypic variation was accounted for the region located between RM311 and RM6100. Using the marker-trait segregation data derived from analysis of both the mapping populations, a local linkage map of the genomic region around Rf-4, a major fertility restoration locus on Chromosome 10 was constructed, and RM6100 was observed to be very close to the gene at a distance of 1.2 cM. The accuracy of the marker RM6100 in predicting fertility restoration was validated in 21 restorers and 18 maintainers. RM6100 amplified the Rf-4 linked allele in a majority of the restorers with a selection accuracy of 94.87%. Through the present study, we have established the usefulness of the marker RM6100 in marker-assisted selection for fertility restoration in segregating populations and identification of restorers while screening rice germplasm for their fertility restoration ability.     

Rht1(B. dw), an alternative allelic variant for breeding semi-dwarf wheat varieties

The vast majority of the world's acreage of semi-dwarf wheat varieties is at present cultivated with varieties carrying one of two genetically similar dwarfing genes, Rht1 and Rht2, derived from the Japanese variety ‘Norin 10’. Near-isogenic lines have been developed and tested to determine the breeding potential of an allelic variant of Rht1, designated Rht1(B. dw). Following its introduction into four varietal backgrounds, Rht1 (B. dw) was seen to reduce height by around 25%, to increase the number of grains setting in spikelets and ears by around 20%, to reduce grain weight by 10%, and to increase yields of plants grown under spaced or drilled conditions by about 8%. When compared to the commercially utilized Rht1 allele, as near isogenic lines in a ‘Mercia’ varietal background, Rht1 (B. dw) gives a significantly greater reduction in plant height, a greater increase in spikelet and ear fertilities, slightly less reduction in 1000-grain weight, and significantly higher spikelet, ear and plot yields. If these results are repeatable in other varietal backgrounds, over seasons, and under differing environmental conditions, Rht1 (B. dw) should have considerable commercial potential as an alternative allele for producing shorter-than-average, high-yielding, semi-dwarf wheat varieties.