Random amplified polymorphic DNA analyses of cytoplasmic male sterile and male fertile Pigeonpea (Cajanus cajan (L.) Millsp.)

Randomly amplified polymorphic DNA (RAPD) markers were used for the identification of two pigeonpea cytoplasmic genic male sterile (CMS) lines derived from crosses between the wild (Cajanus scarabaeoides & C. sericeus) and the cultivated species of Cajanus cajan. The male sterile (A) line and its maintainer (B) line could be easily differentiated with certain random primers. The two male sterile (288 A and 67 A)systems are based on C. scarabaeoidesand the other is based on C. sericeus could also be differentiated. Amplification product of 600bp amplified by primer OPC-11 was observed in both the cytoplasmic male sterile lines (288 A and 67A), which was absent in the maintainer lines (288 B and 67 B) and the putative R-line (TRR 5 and TRR 6). CMS lines, putative R lines, other cultivars and wild species under the study could be easily distinguished with the help of different primers. Dendrogram constructed based on the similarity index showed that considerable genetic variation exist sbetween CMS lines, two putative R line and wild species studied. This revised version was published online in July 2006 with corrections to the Cover Date.     

木豆CGMS杂交种生产中的传粉昆虫

木豆是世界上唯一的木本食用豆类作物,具有较强的抗旱性。木豆杂交制种需昆虫传粉,制种地传粉昆虫的种类及数量是制种产量的决定性因素。而对木豆杂交制种中传粉昆虫与种子产量关系的研究极为少见。国内尚无传粉昆虫种类、数量及木豆杂交种制种产量的任何研究报道。本文调查了云南省元谋县CGMS木豆[Cajanus cajan (L.) Millspaugh]杂交种ICPH2671制种田访花昆虫种类、数量、访花频次及杂交种制种产量等。结果表明在云南元谋,木豆访花昆虫共有5目15科25种,其中主要5种传粉媒介为绒切叶蜂(Megachile velutina)、切叶蜂属一种(Megachile sp5)、圆柄木蜂(Xylocopa tenuiscapa)、蜜蜂亚科一种(Apinaesp)和切叶蜂属一种(Megachile sp2)。木豆盛花期,制种亲本不育系每个分枝昆虫访花频次为每10 min 2.8次,而恢复系植株上的访花频次为每10 min 5.2次,表明昆虫对恢复系花朵有明显偏好;在前述昆虫访花频次下,不育系单株干籽粒产量达383.7 g,恢复系单株产量为357.0 g,二者无显著差异,说明这样的昆虫访问频次已能满足杂交制种中将花粉由恢复系传至不育系的需要。     

Molecular characterization of fertile and sterile cytoplasms in Beta spp.

Mitochondrial DNA (mtDNA) from sugar beet carrying fertile (F) and male sterile (CMS) cytoplasms, and from male sterile accession of Beta maritima collected in Brittany (France) were characterized and compared by restriction fragment length polymorphism (RFLP) and Southern hybridization with coxII. The F and CMS cytoplasms could be clearly distinguished from each other by RFLP when XhoI, EcoRI and BamHI endonucleases were used. Southern hybridization with the coxII gene provided further evidence that mitochondrial genome organization differs between fertile and sterile plants. All cytoplasmic male sterile lines from different breeding stations showed the same restriction and hybridization patterns, which confirms the uniformity of mitochondrial genomes within the materials used for hybrid seed production in several European countries. No visible differences were found between the maintainer lines studied. However, comparisons of XhoI restriction profiles of mtDNA from maintainer lines and from fertile monogerm populations revealed slight differences, which were reflected by the appearance of a unique 0.9 kb fragment in the latter. Analysis of mtDNA from male sterile plants of the wild beet B. maritima showed different restriction and hybridization patterns in comparison with normal and sterile sugar beet cytoplasms. This shows the unique nature of cytoplasmic male sterility in this species.     

Maternal inheritance of male sterility in the progeny of a natural hybrid between Cajanus lineatus and C. cajan

Adoption of pigeonpea hybrids in central and southern India is showing high impact with on‐farm yield advantages of >30%. The hybrid pigeonpea technology, the first in any legume crop, is based on a cytoplasmic‐nuclear male‐sterility (CMS) system. For a long‐term sustainability of hybrid programme, it is imperative that both nuclear diversity and cytoplasmic diversity are maintained among hybrid parents. In this context, a continuous search for new CMS‐inducing cytoplasms is necessary. This paper reports detection of maternal inheritance of male sterility in the progeny derived from a natural hybrid between a wild relative [Cajanus lineatus (W. & A.) Maesen comb. nov.] of pigeonpea and an unknown pigeonpea [Cajanus cajan (L.) Millsp.] genotype. In the present study, the male sterility was maintained up to BC₇F₁ generation by an advanced breeding pigeonpea line ICPL 99044. This male sterility inducing cytoplasm of C. lineatus was tagged as A₆. In future, this CMS genetic stock can be used to develop a range of new pigeonpea hybrids with high yield and adaptation.     

Chromosome 1D as a possible location of a gene (s) controlling variation between wheat (Triticum aestivum L.) varieties for carbon isotope discrimination (Δ) under water-stress conditions

Cytoplasmic male-sterility (CMS) in pigeonpea has been reported when some wild relatives of pigeonpea were crossed as the female parent with cultivated types as the male parent. In this paper we report a new source of CMS developed by using the cultivated pigeonpea as the female parent and one of its wild relative Cajanus acutifolius as the pollen donor. This is the first report in pigeonpea where CMS has been developed using the cytoplasm of cultivated pigeonpea. Several pure line cultivars of pigeonpea restored pollen fertility whereas cv. HPL 24 partially maintained male-sterility. The wild species C. acutifolius used as one of the parents, maintained complete sterility. Cytological analysis revealed that both in male-sterile as well as the fertile floral buds, meiosis proceeded normally till the tetrad stage. However in the male-sterile genotypes during the formation of tetrads, the pollen mother cell (PMC) wall did not dissolve to release the tetrads unlike in the fertile genotypes and this major event was found to be responsible for male-sterility.     

Diversity in selected wild and cultivated species of pigeonpea using RFLP of mtDNA

Diversity in 28 accessions representing 12 species of the genus, Cajanus arranged in 6 sections including 5 accessions of the cultivated species, C. cajan, and 4 species of the genus Rhyncosia available in the germplasm collection at ICRISAT was assessed using RFLP with maize mtDNA probes. Cluster analysis of the Southern blot hybridization data with 3 restriction enzymes – 3 probe combinations placed the genus Rhyncosia in a major group well separated from all the species belonging to the genus Cajanus. Within the genus Cajanus, the 4 accessions of C. platycarpus belonging to section Rhynchosoides formed a separate group in contrast to those in other sections of pigeonpea. In the section, Cajanus all the 5 accessions of C. cajan were grouped together and C. cajanifolius belonging to the same section was in a subgroup by itself closer to the main group. The four accessions of C. scarabaeoides, were together and the other species belonging to section Cantharospermum were in different subgroups. The intra-specific variation was seen even within accessions of certain pigeonpea wild species such as C. scarabaeoides, C. platycarpus, C. acutifolius, and even the cultivated species of C. cajan. This study suggests that RFLP of mtDNA can be used for the diversity analysis of pigeonpea and it gives some indications on the maternal lineage among the species. The variations in the mitochondrial DNA hybridization patterns also suggest the extensive rearrangement of the organelle genome among the Cajanus species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Production of hybrids between Cajanus acutifolius and C. cajan

There are many wild species of pigeonpea which are endemic to Australia. These wild species are cross incompatible with cultivated species of Indian origin. Cajanus acutifolius is one such species which does not easily cross with cultivated pigeonpea. Interspecific pollinations lead to hybrid seeds which were semi-shrivelled. Very few seeds germinated to give rise to F1 plants. Backcrossing the hybrid plants to C. cajan, the male parent, gave rise to aborting seeds which did not germinate in vivo hence BC1 plants are obtained after saving the aborting embryos in vitro. BC₁ plants showed normal meiotic pairing, but had low pollen fertility. The reasons for embryo abortion and low pollen fertility in spite of normal meiosis could be due to the effect of wild species cytoplasm. This revised version was published online in August 2006 with corrections to the Cover Date.     

A cytoplasmic-nuclear male-sterility system derived from a cross between <Emphasis Type="Italic">Cajanus cajanifolius</Emphasis> and <Emphasis Type="Italic">Cajanus cajan</Emphasis>

Cytoplasmic-nuclear male-sterility is an important biological tool, which has been used by plant breeders to increase yields in cross-pollinated cereals and vegetables by commercial exploitation of the phenomenon of hybrid vigor. In legumes, no such example exists due to the absence of an economic way of mass pollen transfer from male to female parent. Pigeonpea [Cajanus cajan (L.) Millsp.], however, is a different legume where a moderate level of insect-aided natural out-crossing (25–70%) exists and it can be used to produce commercial hybrid cultivars, if an efficient and stable cytoplasmic-nuclear male-sterility (CMS) system is available. This paper reports the development of a stable CMS system (ICP 2039A), derived from an inter-specific hybrid of Cajanus cajanifolius, a wild relative of pigeonpea, with a cultivar ICP 11501. Using this genetic material, designated as the A₄ cytoplasm, a number of fertility restorers and maintainers have been developed. The best short-duration experimental pigeonpea hybrid ICPH 2470 produced 3205 kg ha⁻¹ grain yield in 125 days, exhibiting 77.5% advantage over the control cultivar UPAS 120. At present, all the important biological systems necessary for a successful commercial hybrid breeding program are available in pigeonpea and the package of this technology has been adopted by private seed sector in India for the production and marketing of hybrid varieties.     

小麦质核互作型雄性不育系线粒体DNA变异性的RAPD分析(英文)

在杂交小麦选育中,为了更好地利用细胞质雄性不育性,研究与不断挖掘新的不育细胞质类型及其对应育性恢复基因以改良现有不育资源至关重要.基于这一目的,本文对具有山羊草属细胞质的四类不育系线粒体DNA(mtDNA)进行了RAPD分析,分别比较了具有同一细胞质背景的山羊草、雄性不育系,以及该类不育系与恢复系组配的可育杂种F1的mtDNA的变异性.结果显示,供试山羊草与其对应细胞质雄性不育系在mtDNA上存在明显多态性,表明不育系在质核互作的影响下很可能已导致mtDNA发生变异:而不育系与对应的可育杂种F1在mtDNA上也存在多态性,同样表明育性恢复核基因对不育系进行育性恢复的过程中亦可能引起mtDNA发生相应变异;mtDNA变异很可能涉及到不育系育性本质的改变.     

木豆种质资源AFLP标记遗传多样性分析

利用AFLP分子标记技术,以木豆野生种(Cajanus scarabaeiodes)为对照,对来自印度、中国、非洲和美洲的139份栽培木豆(Cajanus cajan)种质遗传多样性进行研究。17对引物共扩增出502条清晰可辨的带,其中多态性带494条。遗传相似性系数UPGMA聚类图,揭示木豆种内存在足够的遗传多样性,可用于种质资源的准确鉴别与分类。借助遗传距离的ME进化树又将参试种质分成8个组群。分析结果表明,印度地方资源遗传多样性广泛,育成品种遗传基础广阔;非洲、美洲资源遗传多样性较丰富,且与印度资源的亲缘关系较密切。中国地方资源遗传多样性独特,且与印度、非洲、美洲栽培木豆资源亲缘关系不明显。研究结果基本支持印度木豆起源和多样性中心、非洲次生多样性中心的观点,并提出中国次生起源中心和遗传多样性中心假说。研究还发现,木豆核质互作雄性不育系(A)和保持系(B)间存在独特的AFLP谱带差异,可能有助于分子标记辅助的木豆杂种优势利用和杂交种培育。     

Production of hybrids betweenCajanus platycarpus andCajanus cajan

Summary Cajanus platycarpus a wild species of pigeonpea incompatible with the cultigen has many desirable characters important for the improvement of cultivated pigeonpea. In the present study, barriers to hybridization were studied and were identified as post-zygotic. Efficient embryo rescue techniques were developed. As a result, hybrids were produced. Morphological and cytological studies as well as esterase isozyme band pattern confirmed the hybrid nature of the plants. The F₁ hybrids were completely pollen sterile. Meiotic studies were carried out to check for the cause of pollen sterility.Abbreviations MS Murashige & Skoog's (1962) medium - BAP 6-Benzylaminopurine - IBA Indole-3-butyric Acid - NAA Naphthaleneacetic Acid     

Chloroplast-DNA variation in the wild and cultivated olives (Olea europaea L.) of Morocco

The male sterile plants that segregated in a BC₅F₂ of `C. sericeus × C. cajan var. TT-5' population were maintained by sib mating. The male sterile plants were crossed with ICPL-85012.Approximately 50% of the F₁ plants were sterile. F₂ plants derived from the fertile F₁ plants did not segregate for male sterility. The reciprocal hybrid i.e. ICPL-85012 × Fertile derivatives from C. sericeus × TT-5, did not express male sterility. However, among the 12 F₂ plant to row progenies, two segregated 25% male sterile plants and remaining 10 did not segregate. The segregation pattern in subsequent progenies revealed that the sterility was under control of a single recessive allele. Studies on the backcross and their BC₁F₂ and BC₁F₃progenies revealed another sterility gene which was found to be dominant in inheritance. This paper shows that what was thought to be cytoplasmic male sterility from C. sericeus cytoplasm is actually a single dominant gene possibly acting in concert with a single recessive gene to mimic cytoplasmic male sterility. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cajanus platycarpus (Benth.) Maesen as the donor of new pigeonpea cytoplasmic male sterile (CMS) system

Cajanus platycarpus, a distantly related wild species, was successfully crossed with cultivated pigeonpea using embryo rescue and tissue culture techniques. Advance generation lines showed a range of desirable characters including cytoplasmic male sterility. A range of pigeonpea cultivars restored fertility and was maintained by a few lines including cultivar ICPL 85010. Clasmogamous flowers were observed in the male sterile lines. In such flowers anthers did not form di-adlephous bundle. Cytological analysis revealed that meiosis proceeded normally till the tetrad stage in those anthers with pollen grains. After which many of the pollen grains turned sterile. In the anthers with pollen grains, dehiscence was not observed, thus creating functional sterility. In many other anthers, pollen mother cells (PMCs) were not formed at all, giving rise to sepalous anthers. In conclusion two mechanisms of male sterility existed, one was premeiotic, where PMCs did not form and in the second, although PMCs gave rise to pollen grains, they were either partially or totally sterile accompanied by non-dehiscence of anther wall.     

Obtaining an Ogura-type CMS line from asymmetrical protoplast fusion between cabbage (fertile) and radish (fertile)

Cytoplasmic male sterile (CMS) plants were obtained by asymmetrical protoplast fusion between red cabbage (fertile) and normal radish (fertile). The CMS plants showed restriction fragment length polymorphism (RFLP) patterns of mitochondrial (mt) DNA that were different from those of both parental lines. PCR analysis of mtDNA of the CMS plants and though Southern blot analysis showed that the mtDNA of the CMS line had the characteristics of the ogura CMS type. The results suggested that the orf138 gene and the ogura type atp6 gene are present in normal fertile cabbage and radish at a low copy level. This revised version was published online in July 2006 with corrections to the Cover Date.     

The Influence of Genotype–Environment Interaction on the Grain Yields of 10 Pigeonpea Cultivars Grown in Kenya

To study the importance of the effects of genotype–environment interactions on the yield of pigeonpea ( Cajanus cajan L. Millsp.), 10 early-maturing genotypes were grown in a randomized complete block design with three replications in a total of seven environments spread over five regions of Kenya between 1987 and 1988. Results indicated the presence of a substantial genotype–environment interaction effect on grain yield. The observed significant genotype–environment interaction effect is discussed in relation to its importance in pigeonpea grain yield evaluation studies. It is noted that the best genotype in one environment is not always so in other environments. Results from regression analysis indicated that this method of analysis is appropriate for describing the response of pigeonpea genotypes grown in a number of locations. Analysis of variance showed significant additive and multiplicative genotype–environment interaction effects. Only the first interaction principal component axis (IPCA) was found to be important in describing the multiplicative interaction effects. The additive main effects and multiplicative effects (AMMI) model allowed the partitioning of interaction variance into agronomically important sources (genotype groups), and the specific genotype × environment patterns that are the basis of these sources of variance were examined.     

Genetics of fertility restoration in A2-based cytoplasmic genetic male sterility system in pigeonpea (Cajanus cajan)

The three short duration cytoplasmic genetic male sterility (CGMS) hybrids developed using A₂ (Cajanus scarabeoides) cytoplasm-based male sterile lines (CORG 990047A, CORG 990052A and CORG 7A) and the restorer inbred AK 261322 and their segregating populations (F₂ and BC₁F₁) were subjected to the study of inheritance of fertility restoration in pigeonpea. The fertility restoration was studied based on three different criteria, namely, anther colour, pollen grain fertility and pollen grain morphology and staining. The F₂ and BC₁F₁ populations of the three crosses, namely, CORG 990047A × AK 261322, CORG 990052A × AK 261322 and CORG 7A × AK 261322, segregated in the ratio of 3:1 and 1:1, for anther colour (yellow:pale yellow), pollen grain fertility (fertile:sterile) and for pollen grain morphology and staining. The above study confirmed that the trait fertility restoration was controlled by single dominant gene. This finding can be utilized for the identification of potential restorers, which can be further used in the development of CGMS-based hybrids in pigeonpea.     

水稻野败不育系与保持系线粒体DNA限制酶酶切图谱分析

采用限制酶酶切片段长度多态性（ＲＦＬＰ）分析广泛比较了水稻野败不育系及其籼型保持系线粒体ＤＮＡ酶切图谱。野败不育系与籼型保持系线粒体ＤＮＡ在组成上存在十分明显的差异。根据具有相同迁移位置与大小的片段比例判断，这２种不同细胞质线粒体ＤＮＡ顺序同源性约为３０％。分子杂交结果显示，编码功能蛋白的基因及其邻近顺序存在在相当程度的保守性。本文并对线粒体ＤＮＡ的组成变异与细胞质雄性不育的关系进行了分析与讨论。     

细胞质雄性不育系在大豆常规育种中的应用研究

提高育种效率是育种家追求的育种手段之一,大豆细胞核雄性不育系已被应用于轮回选择育种中,但细胞质雄性不育系能否应用于常规大豆育种程序还未曾有报导。此研究以13个细胞质雄性不育系和9个恢复系配制15个杂交组合,利用苜蓿切叶蜂授粉,通过对其后代的选择与鉴定,选育出4个高产、优质大豆新品系。探讨了该方法的育种效率、组合选配与后代选择问题以及杂种优势与后代表现的关系。研究结果显示,该方法能显著提高育种效率,为细胞质雄性不育系在常规大豆育种中的应用开辟了新的途径。     

Introgression of Cajanus platycarpus genome into cultivated pigeonpea, C. cajan

Summary Cajanus platycarpus, an incompatible wild species from the tertiary gene pool of pigeonpea (C. cajan (L.) Millspaugh), has many desirable characteristics for the improvement of cultivated varieties. To necessitate such transfers, embryo rescue techniques were used to obtain F₁ hybrids. The F₁ hybrids were treated with colchicine to obtain tetraploid hybrids, that were selfed to obtain F₂, F₃ and F₄ progenies. All of the hybrids and subsequent progenies had an intermediate morphology between the two parents. Backcrossing of the tetraploid hybrids with cultivated pigeonpea was not possible given embryo abortion, with smaller aborted embryos than those obtained in the F₀ parental cross.As a route of introgression, diploid F₁ hybrids were backcrossed with cultivated pigeonpea and BC₁ progeny obtained by in vitro culture of aborting embryos. BC₂ plants were obtained by normal, mature seed germination. Although embryo rescue techniques had to be used to obtain F₁ and BC₁ plants, it was possible to produce BC₂ and subsequent generations through direct mature seed. Every backcross to cultivated pigeonpea increased pollen fertility and the formation of mature seeds.Special project assistant till December, 2003.     

Identification of the Putative Cytoplasmic Donor of a CMS System in Brassica juncea

Chloroplast (cp) and mitochondrial (mt) DNA restriction profiles of a cytoplasmic male sterile (CMS) line of Brassica juncea and its maintainer line were compared and found to be markedly different. Comparison of cpDNA restriction profiles of fifty different species of genus Brassica and some allied genera showed that the cpDNA profiles of CMS lines were similar to that of B. tournefortii for twenty different restriction endonucleases. This CMS system is thus not of spontaneous origin as reported earlier, but is alloplasmic in nature. Comparison of restriction profiles of mtDNA of B. tournefortii and CMS lines revealed some differences which might either be due to changes in DNA pattern during the transfer, or, due to the cytoplasm coming from a B. tournefortii line different from the one used in this study.