Cajanus Scarabaeoides Research Articles

Genetic Diversity Studies and Screening for Fusarium Wilt (Fusarium udum Butler) Resistance in Wild Pigeonpea Accessions, Cajanus scarabaeoides (L.) Thouars

Wild relatives play an important role in the genetic improvement of cultivated crops. Cajanus scarabaeoides (L.) Thouars is one of the important wild species, which possess several desirable traits. Investigations on 67 accessions of C. scarabaeoides and 3 cultivated varieties for total soluble seed protein profiles using Sodium Dodecyl Sulphate-Poly Acryl amide Gel Electrophoresis resolved the protein bands ranged from 4–11. Based on the presence or absence of bands, similarity values were calculated and dendrogram was constructed using Unweighted Pair Group Method with Arithmetic Mean analysis. The dendrogram formed two main clusters. The first main cluster consisted of three cultivated varieties and 55 accessions of C. scarabaeoides and the second main cluster comprised of remaining 12 accessions of C. scarabaeoides. Similarity percentage ranged from 38.5 to 100. ICP15728 is the lone genotype which is distinctly different from remaining genotypes formed separate sub-cluster in the second main cluster with 38.5% similarity. Thirty eight accessions of C. scarabaeoides were screened against Fusarium wilt under glass house conditions. Of the 38 accessions screened seven accessions (ICP12707, ICP15689, ICP15692, ICP15732, ICP15744, ICP15748 and ICP15754) showed resistance to Fusarium wilt. Therefore, these accessions could be utilized in breeding programmes to increase the levels of resistance to Fusarium wilt in the pigeonpea cultivars, after further testing.

Molecular characterization of cultivated and wild <italic>Cajanus</italic> species using Simple Sequence Repeat markers

Twenty pigeonpea specific SSR markers were used to test the molecular diversity among 21 cultivated genotypes and three wild species. Twelve markers were monomorphic and twenty two alleles were produced by eight polymorphic markers. Ten markers were transferrable among the cultivated and wild species. The PIC value of the markers ranged from 0.081(PGM 3 and PGM 82) to 0.603 (PGM 106). The average PIC value of the markers was 0.30. Based on the dendrogram constructed using the dissimilarity values, 24 accessions were grouped into two main clusters. Main cluster I comprises all the cultivated genotypes, and wild species <italic>viz., Cajanus scarabaeoides</italic> and <italic>Cajanus cajaniolius. Cajanus sericeus</italic> formed the separate cluster (Main cluster II). Within the main cluster I, seven sub-clusters were observed. <italic>Cajanus scarabaeoides</italic> and <italic>Cajanus cajaniolius</italic> formed a separate sub-cluster within the main cluster I. Most of the diversity was observed in between the wild species and cultivated genotypes. There is no significant molecular diversity among the cultivated genotypes. Such a narrow genetic base is likely to represent a serious impediment to breeding programs in pigeonpea. This study suggested the need for broadening the genetic base of cultivated genotypes to improve the productivity of the crop.

Development of dominant sequence characterized amplified region (SCAR) marker linked with plume moth (Exelastis atomosa Walsingham 1886) resistance in pigeon-pea

The mode of gene action governing resistance to plume moth (Exelastis atomosa Walsingham 1886) derived from pigeon-pea (Cajanus scarabaeoides (L.) Thouars) accession ICPW-94 has been determined and the resistance alleles have been designated as PPM1. The progenies of F2 population and F3 families derived from an interspecific cross C. cajan (L.) Huth ('ICP-26') x C. scarabaeoides (accession ICPW-94) revealed monogenic gene action for resistance to plume moth, and the dominant control by single locus or cluster of tightly linked alleles. Bulked segregant analysis (BSA) of 116 F2 progenies by using 143 parental polymorphic RAPD primers could identify a fragment OPA09910 associated with plume moth resistance in coupling phase of linkage. Further single plant analysis of the 116 F2 mapping population revealed OPA09910 was linked to PPMi locus conferring host resistance to plume moth with recombination fraction (rf) value of 0.125 (12.7 cM of Kosambi function). The resistance specific fragment OPA09910 was cloned, sequenced and converted into a sequence characterized amplified region (SCAR) marker, SCOPA09942, which was also closely associated (10.3 cM) with the locus PPMl with rf value 0.102. BLAST analysis with pigeon-pea genome sequence also confirmed its occurrence in CcLG02 (Scafseq.LG_V5.0fa) and contig 01597 (AFSP01.fsa1). This SCAR marker showed reasonable screening efficiency in the F2, F3, and BC1F1 lines, thus it can be used as genetic handle in marker-assisted introgression of the genomic fragment conferring plume moth resistance and screening of breeding lines in pigeon-pea.

A Study on Comparative Fertility Restoration in A2 and A4 Cytoplasms and Its Implication in Breeding Hybrid Pigeonpea [Cajanus cajan (L.) Millspaugh

Exploitation of hybrid vigour has been visualized as the most efficient option for increasing productivity in pigeonpea [Cajanus cajan (L.) Millspaugh]. Cytoplasms from various wild relatives of pigeonpea have been transferred to develop CMS lines in the background of cultivated pigeonpea. However, A2 (Cajanus scarabaeoides) and A4 (Cajanus cajanifolius) cytoplasms have been utilized most frequently. In order to study fertility restoration efficiency in F1 hybrids having either A2 or A4 cytoplasms, an experiment was conducted at the Indian Institute of Pulses Research (IIPR), Kanpur during 2008-2012. Four CMS lines namely Hy4A, H28A (each with A2 cytoplasm), ICP 2039A and ICP 2043A (both with A4 cytoplasm) were crossed with ten genotypes/restorers of long duration pigeonpea for two years. The F1 hybrids so-obtained were assessed in the succeeding years for pollen fertility and pod setting. All the pollinators except IPA 203 restored fertility in F1 hybrids derived from ICP 2039A and ICP 2043A (both having A4 cytoplasm). However, none of the restorers were effective in restoring fertility in hybrids derived from Hy4A and H28A (each with A2 cytoplasm). This could be ascribed to undesirable linkage drag still present in these two CMS lines having A2 cytoplasm. The F2 progenies derived from 4 hybrids (ICP 2039A × NA-1, ICP 2039A × Bahar, ICP 2043A × NA-1 and ICP 2043A × Bahar) segregated approximately into 3 fertile: 1 sterile plants. However, 2 F2 progenies having Pusa 9 as the restorer revealed approximately 15 fertile:1 sterile ratio. Thus monogenic and digenic duplicate gene action with complete dominance for fertility restoration was observed in F1 hybrids derived from CMS lines having A4 cytoplasm. F3 progenies from individual F2 plants of these crosses also confirmed the same pattern of fertility restoration. This study indicated that CMS lines based on A4 cytoplasm would be more desirable as these might have more number of restorers compared to those having A2 cytoplasm.

Yield factor analysis in F4and F5progenies derived from interspecific hybridization between cultivated and wild pigeonpea [Cajanus cajan(L.) Millsp

A study on 81 F4 and F5 progenies derived from the interspecific crosses between cultivated types viz. UPAS 120, Pant A 134 and ICPL 84023 and wild types viz. Cajanus cajanifolius, Cajanus scarabaeoides and Cajanus acutifolius revealed significant differences among all the treatments. High estimates of PCV and GCV were obtained for seed yield per plant and number of pods per plant. Pods per plant revealed wide spectrum of variability in both F4 and F5 progenies. In general, the genotypic correlation coefficients were higher in magnitude than the corresponding phenotypic correlation coefficients. During both the cropping seasons, number of pods per plant and seed yield per plant showed high / moderate estimates of heritability coupled with high genetic advance indicating the preponderance of additive gene effects. Seed yield per plant showed significant positive correlation with number of pods per plant and plant height in both the generations. Path coefficient analysis revealed that in both the generations number of pods per plant, 100 seed weight and number of seeds per pod contributed maximum towards the total yield.

Pigeonpea genotypes influence parasitization preference and survival and development of the Helicoverpa armigera larval parasitoid, Campoletis chlorideae.

Studies were undertaken to identify pigeonpea, Cajanus cajan (L.) Millspaugh and the wild relative of pigeonpea, Cajanus scarabaeoides (L.) (accession ICPW 125,) genotypes that are hospitable to the pod borer, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) larval parasitoid, Campoletis chlorideae Uchida (Hymenoptera: Ichneumonidae) for the management of this pest in pigeonpea based cropping systems. Percentage parasitization of the H. armigera larvae by the C. chlorideae females was greater under no-choice conditions than under multi-choice conditions because of forced parasitization under no-choice conditions. Lowest parasitization was recorded on the wild relative, ICPW 125, which may be due to long nonglandular hairs and low survival of H. armigera larvae. Parasitization of H. armigera larvae was greater under no-choice, dual-choice and/or multi-choice conditions on ICPL 87, ICPL 87119 and ICPL 87091, which are susceptible to H. armigera, than on the pod borer-resistant genotypes ICPL 332WR, ICPL 84060 and ICPB 2042; while survival and development of the parasitoid was better on H. armigera larvae fed on ICPL 87, ICPL 87119, LRG 41, ICP 7035 and ICPL 87091 than on ICPL 332WR, ICPL 84060, ICPB 2042 and ICPW 125. The genotypes ICPL 87, ICPL 87119, LRG 42 and ICPL 87091 that are hospitable to C. chloridae, are better suited for use in integrated pest management to minimize the losses due to H. armigera in pigeonpea.

Host shift capability of a specialist seed predator of an invasive plant: roles of competition, population genetics and plant chemistry

Acanthoscelides macrophthalmus is a seed predator that has become widely distributed along with its native host, Leucaena leucocephala (Mimosoideae), which is a neotropical leguminous tree and one of the most invasive plants worldwide. Previous studies revealed that A. macrophthalmus is able to host-shift to several mimosoid species. Here, we aim to test the host-shift potential to other mimosoid and non-mimosoid plants and possible roles of interspecific competition, genetic background, and plant chemistry in host-shift. First, we found that A. macrophthalmus predator completed development on two new hosts: pigeon pea Cajanus cajan and Cajanus scarabaeoides (Faboideae), by rearing from seeds collected in South/Southeast Asia and Hawaii. In contrast, in most regions, both Cajanus species were infested only by other beetle species. Second, we performed no-choice tests using 11 leguminous plants, covering all three subfamilies as potential hosts, including the two new hosts. A Taiwanese A. macrophthalmus population reared in the laboratory on Leucaena did not deposit eggs on any of the seeds of each tested species. To compare host-shift responses between populations, we also used a Hawaiian A. macrophthalmus population that had completed its development on freshly collected Leucaena seeds from the field. This population deposited eggs onto and hatching larvae burrowed into C. cajan seeds, although none developed beyond the larval stage. Third, the surface chemical composition of seed-pods of L. leucocephala and the two Cajanus species was dissimilar, although that of seeds was highly similar. Finally, all of the host-shifting A. macrophthalmus populations shared the same haplotypic group.

Novel genic microsatellite markers from Cajanus scarabaeoides and their comparative efficiency in revealing genetic diversity in pigeonpea

Paucity of molecular markers is hindering molecular breeding programmes for genetic improvement in pigeonpea, which is considered to be among the richest source of dietary protein in Asia and Africa. At the time of the start of this study, only 156 microsatellite markers were available in pigeonpea (Burns et al. 2001; Odeny et al. 2007, 2009). Recently with the publication of draft genome sequence and deep transcriptome studies, the stage has been set to enrich genomic resources to aid molecular breeding in pigeonpea (Dutta et al. 2011; Singh et al. 2012; Varshney et al. 2012). Genic microsatellites or EST-SSRs (simple sequence repeats) derived from expressed sequence tags (ESTs) are useful because these are inexpensive to develop, represent transcribed genes, and often a putative function can be assigned to them. Compared with genomic sequences, genic SSRs have several advantages as genetic markers. First, if an EST marker is found to be genetically associated with a trait of interest, it may represent the gene affecting the trait directly (Chen et al. 2001; Thiel et al. 2003). Therefore, EST-derived markers can provide opportunities for gene discovery and enhance the role of genetic markers by assaying variation in transcribed and known-function genes. Second, EST-derived

Large-Scale Development of Cost-Effective Single-Nucleotide Polymorphism Marker Assays for Genetic Mapping in Pigeonpea and Comparative Mapping in Legumes

Single-nucleotide polymorphisms (SNPs, >2000) were discovered by using RNA-seq and allele-specific sequencing approaches in pigeonpea (Cajanus cajan). For making the SNP genotyping cost-effective, successful competitive allele-specific polymerase chain reaction (KASPar) assays were developed for 1616 SNPs and referred to as PKAMs (pigeonpea KASPar assay markers). Screening of PKAMs on 24 genotypes [23 from cultivated species and 1 wild species (Cajanus scarabaeoides)] defined a set of 1154 polymorphic markers (77.4%) with a polymorphism information content (PIC) value from 0.04 to 0.38. One thousand and ninety-four PKAMs showed polymorphisms between parental lines of the reference mapping population (C. cajan ICP 28 × C. scarabaeoides ICPW 94). By using high-quality marker genotyping data on 167 F2 lines from the population, a comprehensive genetic map comprising 875 PKAMs with an average inter-marker distance of 1.11 cM was developed. Previously mapped 35 simple sequence repeat markers were integrated into the PKAM map and an integrated genetic map of 996.21 cM was constructed. Mapped PKAMs showed a higher degree of synteny with the genome of Glycine max followed by Medicago truncatula and Lotus japonicus and least with Vigna unguiculata. These PKAMs will be useful for genetics research and breeding applications in pigeonpea and for utilizing genome information from other legume species.

Cyto-morphological and molecular characterization of Cajanus cajan × C. scarabaeoides F1 hybrid

Cajanus scarabaeoides has many desirable agro- nomic traits that deserve attention for introgression into pigeonpea cultivars. An inter-specific F1 hybrid was raised using C. cajan (ICP-26) as seed parent and C. scara- baeoides (ICPW-94) as pollen parent. Hybridity of the F1 plant was affirmed by analyzing the morphological attrib- utes, such as seed coat color, leaf shape, brown striations in the petal, pod features, and intermediate branching type etc. Most of the pollen mother cells (PMC) in the F1 hybrid have shown regular meiosis, but in some PMCs chromosome heteromorphism was observed for two pairs of bivalents during diakinesis, leading to precocious separation of biva- lents (62.81 %) at metaphase-I, and bridge formation (7.44 %) and laggards (2.48 %) at anaphase-I. SDS-PAGE of two major seed protein fractions (albumin and globulin) detected five polypeptide bands (Mol wt. 40.7, 39.8, 32.4, 23.5 and 19.5 KDa) unique to C. scarabaeoides, the male parent, in the F1 hybrids that evidenced the hybridity of F1 plants. Further, RAPD and ISSR marker analysis revealed the hybrid nature of the putative F1 plant. These analyses showed the inheritance of 28 RAPD and 12 ISSR markers, unique to C. scarabaeoides - the pollen parent, to F1 hybrids. This complementary use of cyto-morphological analysis, and molecular marker based genotyping allowed precise evaluation of the C. cajan×C. scarabaeoides inter- specific hybrid.

The first genetic map of pigeon pea based on diversity arrays technology (DArT) markers

With an objective to develop a genetic map in pigeon pea (Cajanus spp.), a total of 554 diversity arrays technology (DArT) markers showed polymorphism in a pigeon pea F(2) mapping population of 72 progenies derived from an interspecific cross of ICP 28 (Cajanus cajan) and ICPW 94 (Cajanus scarabaeoides). Approximately 13% of markers did not conform to expected segregation ratio. The total number of DArT marker loci segregating in Mendelian manner was 405 with 73.1% (P > 0.001) of DArT markers having unique segregation patterns. Two groups of genetic maps were generated using DArT markers. While the maternal genetic linkage map had 122 unique DArT maternal marker loci, the paternal genetic linkage map has a total of 172 unique DArT paternal marker loci. The length of these two maps covered 270.0 cM and 451.6 cM, respectively. These are the first genetic linkage maps developed for pigeon pea, and this is the first report of genetic mapping in any grain legume using diversity arrays technology.

Pharmacological investigation of Cajanus scarabaeoides in different animal models of diarrhoea.

The different solvent extract of Cajanus scarabaeoides (Family: Fabaceae ) was studied for its antidiarrhoeal properties in experimental diarrhoea, induced by castor oil and magnesium sulphate in rat. At the doses of 500 mg/kg per oral, the methanol extract showed significant antidiarrhoeal activity in castor oil model and magnesium sulphate-induced diarrhea. Where as the petroleum ether extract and chloroform extract did not show any such activity. The methanol extract also significantly, reduced the intestinal transit in charcoal meal test when compared to atropine sulphate (5 mg/kg; i.m.). The results showed that the methanolic extract of Cajanus scarabaeoides have a significant antidiarrhoeal activity and supports its traditional uses in herbal medicine.

Fertility Restoration in Cytoplasmic-Nuclear Male-Sterile Lines Derived from 3 Wild Relatives of Pigeonpea

Three cytoplasmic-nuclear male-sterile (CMS) lines, one each derived from Cajanus sericeus (A(1) cytoplasm), Cajanus scarabaeoides (A(2) cytoplasm), and Cajanus cajanifolius (A(4) cytoplasm), were crossed to 7 pigeonpea (Cajanus cajan (L.) Millsp.) cultivars in a line x tester mating scheme to study the fertility restoration of the CMS lines. Twenty-one F(1) hybrid combinations were planted in unreplicated 3-row plots in 3 environments. There was no effect of environments on the expression of fertility restoration. Pigeonpea cultivar ICPL 129-3 restored fertility in A(1) cytoplasm and maintained male sterility in the other 2 (A(2) and A(4)) cytoplasms. Among crosses involving CMS line (of A(4) cytoplasm) ICPA 2039 one hybrid combination was male-sterile and another male fertile. The remaining 5 combinations segregated for male-fertility (66-84% fertility restoration). Such testers can easily be purified for use in hybrid breeding programs by selfing and single-plant selection for 2-3 generations.

Inheritance of trichomes and resistance to pod borer (Helicoverpa armigera) and their association in interspecific crosses between cultivated pigeonpea (Cajanus cajan) and its wild relative C. scarabaeoides

The legume pod borer, Helicoverpa armigera, is one of the most devastating pests of pigeonpea. High levels of resistance to pod borer have been reported in the wild relative of pigeonpea, Cajanus scarabaeoides. Trichomes (their type, orientation, density and length) and their exudates on pod wall surface play an important role in the ovipositional behavior and host selection process of insect herbivores. They have been widely exploited as an insect defense mechanism in number of crops. In the present investigation, inheritance of resistance to pod borer and different types of trichomes (A, B, C and D) on the pod wall surface in the parents (C. cajan and C. scarabaeoides) and their F1, F2, BC1 (C. cajan × F1), and F3 generations has been studied. Trichomes of the wild parents (high density of the non-glandular trichomes C and D, and glandular trichome B and low density of glandular trichome A) were dominant over the trichome features of C. cajan. A single dominant gene as indicated by the segregation patterns individually will govern each trait in the F2 and backcross generation. Segregation ratio of 3 (resistant): 1 (susceptible) for resistance to pod borer in the F2 generation under field conditions was corroborated with a ratio of 1:1 in the backcross generation, and the ratio of 1 non-segregating (resistant): 2 segregating (3 resistant: 1 susceptible): 1 non-segregating (susceptible) in F3 generation. Similar results were obtained for pod borer resistance under no-choice conditions. Resistance to pod borer and trichomes associated with it (low density of type A trichome and high density of type C) are governed individually by a dominant allele of a single gene in C. scarabaeoides. Following backcrossing, these traits can be transferred from C. scarabaeoides into the cultivated background.

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.