Characterization of chickpea germplasm conserved in the Indian National Genebank and development of a core set using qualitative and quantitative trait data

Chickpea is the third most important pulse crop as a source of dietary protein. Ever-increasing demand in Asian countries calls for breeding superior desi-type varieties, in turn necessitating the availability of characterized germplasm to breeders. The Indian National Genebank,located at the National Bureau of Plant Genetic Resources, New Delhi, conserves 14,651 accessions of chickpea. The entire set was characterized in a single large-scale experiment.High variation was observed for eight quantitative and 12 qualitative agro-morphological traits. Allelic richness procedure was employed to assemble a core set comprising 1103 accessions, 70.0% of which were of Indian origin. Comparable values of total variation explained by the first three principal components in the entire collection(51.1%) and the core(52.4%)together with conservation of nine pairwise r values among quantitative traits in the core collection and a coincidence rate around 99.7% indicated that the chickpea core was indeed an excellent representation of the entire chickpea collection in the National Genebank. The chickpea core exhibited greater diversity than the entire collection in agro-morphological traits, as assessed by higher variance and Shannon–Weaver diversity indices, indicating that the chickpea core maximized the phenotypic diversity available in the Indian chickpea germplasm. The chickpea core, comprising mainly indigenous desi genotypes, is expected to be an excellent resource for chickpea breeders. Information on the chickpea core can be accessed at http://www.nbpgr.ernet.in/pgrportal.

Interaction between seed size and NaCl on germination and early seedling growth of some Turkish cultivars of chickpea(Cicer arietinum L.)

Chickpea is an important food legume crop of Turkey and is largely grown for human consumption on low moisture or salt-affected soils. The objective of the study was to find the effects of NaCl stress at electrical conductivities of 4.5, 8.6, 12.7 and 16.3 dS/m and seed sizes （7, 8 and 9 mm） on germination and early seedling growth of three popular chickpea cultivars （AKN-97, Gokce and Uzunlu-99）. Mean frequency of germination, germination time, germination index, root length, shoot length and seedling fresh weight showed seed size-dependent responses ofcultivars to salt stress. In general, small seeds germinated and grew more rapidly compared to medium and large seeds of the same cultivars against all levels of salt stress, with the best results in cultivar Uzunlu-99. No effect of NaCl treatments was observed on frequency of germination; however, a drastic decrease in early seedling growth was recorded at increased NaC1 concentrations. Regression analysis results showed a significantly positive relationship （P〈0.01） between seed size and mean germination time, whereas a significantly negative relationship was recorded between seed size and germination index, root length, shoot length. Moreover, linear regression values apparently confirmed that increased seed size in each cultivar affected decreased germination index, root and shoot lengths with enhanced mean germination time. Thus, it was concluded that the use of small seeds could considerably reduce the production costs of chickpea in salt-affected soils.

Effects of Baking, Roasting and Frying on Total Polyphenols and Antioxidant Activity in Colored Chickpea Seeds

Chickpea lines with colored testa (seed coat) contain high levels of polyphenolic compounds that exhibit high levels of antioxidant activity. In a previous study, we showed that common processing procedures, such as soaking and cooking, decrease the levels of these bioactive compounds and subsequent overall antioxidant activity. The observed reduction in total phenolic content was due to the movement of polyphenols from the seed coat to the soaking or cooking water. Here, the effects of baking, roasting and frying processes were examined in relation to total phenolic content (TPC), total flavonoid content (TFC) and ferric-reducing ability of plasma antioxidant activity (FRAP AA) of colored chickpea seeds. Baked, fried and roasted colored chickpea seeds had significantly higher levels of TPC, TFC and FRAP AA than regular cream- and beige-colored seeds subjected to the same treatments. In contrast to our previous results with soaking and cooking, baking, frying and roasting retained most of the TPC, TFC and FRAP AA in the final products. Thus, colored chickpeas subjected to these three processing methods might be considered a functional food in addition to its traditional role of providing dietary proteins.

Total Phenolic Content and Antioxidant Activity of Chickpea (<i>Cicer</i><i>arietinum</i>L.) as Affected by Soaking and Cooking Conditions

Chickpea lines with colored testa (seed coat) contain high levels of polyphenolic compounds that exhibit high levels of antioxidant activity. However, common processing procedures, such as soaking and cooking, may decrease the levels of these bioactive compounds and subsequent overall antioxidant activity. Here, the effects of soaking, cooking and steaming processes were examined in relation to total phenolic content (TPC), total flavonoid content (TFC) and ferric reducing ability of plasma antioxidant activity (FRAP AA) of colored chickpea seeds. All processing steps significantly reduced TPC, TFC and FRAP AA in all of the tested chickpea seeds. Nevertheless, soaking the seeds at room temperature (for 22 h) resulted in a smaller decrease in TPC, TFC and FRAP AA than soaking at 60℃ (for 2 h). Moreover, steaming was superior to cooking in terms of conserving polyphenol and antioxidant activity. The observed reduction in TPC was mainly due to leaching of these compounds from the seed coat into the soaking or cooking water. Based on these results, we suggest that soaking at room temperature for 22 h followed by steaming for 1 h is the best method for retaining TPC, TFC and FRAP AA of colored chickpea.

Rheological, Organoleptical and Quality Characteristics of Gluten-Free Rice Cakes Formulated with Sorghum and Germinated Chickpea Flours

Gluten-free bakery products are produced for people suffering from celiac disease due to sensitivity to gluten. So, the aim of this work was to study the effect of partial substitution of rice flour at 20%, 30% and 40% with sorghum and germinated chickpea flours on rheological, physical, sensory properties and staling rate of prepared gluten-free cake. Obtained results showed that germinated chickpea flour had the highest contents of crude protein (23.62%), lipids (4.89%) and crude fibers (5.76%) as compared to other grain flours. Gradual increase in gelatinization temperature and decrease in maximum viscosity and setback were fond of cake batters with increasing the substituting levels of rice flour from 20% to 40% with sorghum or germinated chickpea flours in comparison to rice flour batter. No significant difference was observed in specific volume between rice cake and cake containing 20% of germinated chickpea and sorghum flours which given 2.71, 2.62 and 2.56 cm3/g, respectively. Data also revealed that gluten-free cake lightness (L*) and total intensity significantly increased with 40% of sorghum flour followed by 30 and 20% of sorghum flour. Substitution of 40% rice flour with germinated chickpea flour resulted in significant high in redness (a*), yellowness (b*) and chroma of gluten-free cake being 11.95, 33.72 and 35.77, respectively followed by substituted levels 30% and 20% which attributed to their natural yellow pigments compared with other investigated samples. Overall acceptability of gluten-free cakes was improved after adding 20, 30% of sorghum flour and 20% of germinated chickpea flour which being 90.7, 88.6 and 87.5, respectively compared to rice flour cake (88.4). Rice flour cake was more freshness than wheat flour cake and other treatments during storage duration. After 6 days, rice cake had significant high freshness (306.2%) followed by gluten-free cake samples with 20% of sorghum or germinated chickpea flours which given 294.5% and 289.4%, respectively. Generally, it could be concluded that the quality and nutritional properties of gluten-free rice cakes could be improved when using 20%, 30% of sorghum flour and 20% of germinated chickpea flour as substitution levels of rice flour.

Analysis of Genetic Diversity among Wild and Cultivated Chickpea Genotypes Employing ISSR and RAPD Markers

Chickpea (Cicer airetinum L.) is an important and most preferred food legume in many parts of the world especially in the Indian sub-continent. Molecular analysis of chickpea using DNA technology has been carried out to identify the diverse genetic base of the cultivars for selected preferential introductions as efficient edible elements. A few of these genetic stocks have been documented here to tap their genetic diversity. The status shows that the level of polymorphism in this species is low. Using PCR based markers, marker assisted selection of polymorphy is one of the established techniques. Here, this procedure has been employed to expedite gene/QTL pyramiding in the chickpea. The study presented here includes analysis of 12 germplasms of chickpea. Standard CTAB method has been performed, with certain modifications, to get better intensity and resolution of DNA bands. Extracted DNA, amplified with 41 RAPD and 21 ISSR primers are thereby tested to determine genetic diversity. The presentation discusses the results of chickpea germplasm diversity on the basis of these observations.

Changes in Growth,Photosynthetic Pigments,Cell Viability,Lipid Peroxidation and Antioxidant Defense System in Two Varieties of Chickpea(Cicer arietinum L.)Subjected to Salinity Stress

Salinity is one of the most severe abiotic stresses for crop production.The present study investigates the salinityinduced modulation in growth indicators,morphology and movement of stomata,photosynthetic pigments,activity of carbonic anhydrase as well as nitrate reductase,and antioxidant systems in two varieties of chickpea(Pusa-BG5023,and Pusa-BGD72).On 20^(th) day of sowing,plants were treated with varying levels of NaCl(0,50,100,150 and 200 mM)followed by sampling on 45 days of sowing.Recorded observations on both the varieties reveal that salt stress leads to a significant decline in growth,dry biomass,leaf area,photosynthetic pigments,protein content,stomatal behavior,cell viability,activity of nitrate reductase and carbonic anhydrase with the rise in the concentration of salt.However,quantitatively these changes were less in Pusa-BG5023 as compared to Pusa-BGD72.Furthermore,salinity-induced oxidative stress enhanced malondialdehyde content,superoxide radicals,foliar proline content,and the enzymatic activities of superoxide dismutase,catalase,and peroxidase.The variety Pusa-BGD72 was found more sensitive than Pusa-BG5023 to salt stress.Out of different graded concentrations(50,100,150 and 200 mM)of sodium chloride,50 mM was least toxic,and 200 mM was most damaging.The differential behavior of these two varieties measured in terms of stomatal behavior,cell viability,photosynthetic pigments,and antioxidant defense system can be used as prospective indicators for selection of chickpea plants for salt tolerance and sensitivity.

Research Advances in Functional Constituents of Chickpea

Based on the generalization and summary of the research on the functional constituents of chickpea at home and abroad in recent years,the research advances in some rich functional constituents including isoflavones,proteins and peptides,carbohydrates,saponins and trace elements in chickpeas was reviewed in this paper. It provides a basis for the research,development and utilization of the functional constituents of chickpea in the future.

Research on Fatty Acid and Amino Acid Components of Desi Chickpea Seeds from Xinjiang

[Objective] The paper aimed to determine the fatty acid components and contents,and measure the fractional protein and the amino acid components of Desi chickpea seeds.[Method] The oil of Desi chickpea seeds was extracted from their kernels collected from Xinjiang.The contents and components of fatty acids were tested by GC-MS.The fractional extraction was used to get the proteins of their kernels,and then the content of protein was tested.The amino acid compositions of the proteins were also analyzed.[Result] The fat content of chickpea seeds was as high as up to 18.2%,among which,the unsaturated fatty acids oleic acid and linoleic acid were the major component,accounting for 80% of the total fatty acids;the content of saturated fatty acid was low.The average content of protein was 23.71% in chickpea seeds.Globulin and albumin were the main protein in chickpea seeds,accounting for more than 80% of the total proteins,while glutelin and gliadin contents were low.All the proteins got from chickpea seeds belonged to complete protein,which contained 18 kinds of amino acids.The content of Glu was the highest（15.8%）,and the content of Asp was 10.7%,followed by Arg（9.30%）,Leu（7.56%）,Lys（6.34%）,Phe,Ala,Val and Ile.The contents of Trp,Met and Cys were the least.The chickpea seed was rich in essential amino acids.[Conclusion] The Desi chickpea was a legume crop containing high content of unsaturated fatty acids and rich in amino acids.

Diversity analysis of chickpea (<i>Cicer arietinum</i>L.) germplasm and its implications for conservation and crop breeding

The exploration of genetically variable accessions is the key source of germplasm conservation and potential breeding material for the future. The more diverse group of cultivars provides an ample opportunity to breeders for releasing new and superior varieties, considering their quality traits for direct commercial utilization. In this study, we assessed the genetic diversity of Cicer arietinum 70 accessions from Pakistan and USA using morphological traits, seed protein and molecular markers. Based on four morphological traits, the average coefficient of variation was calculated as 56.8% with significant correlation among yield traits. The analysis revealed that the accessions 1898, 2819, 3022, 3037, 3040, 3043, 3054, 3059 and 3063 were best in performance with a total of 12% environmental error. The cluster analysis based on protein data revealed 50% genetic diversity among accessions. The clustering pattern did not show any grouping that could be attributed to either the geographic distribution or the field performance. For molecular characterization of germplasm 5 PCR based RAPD primers, OPA4, OPA9, OPG13, UBC181 and UBC733b used were found to be polymorphic with 37% genetic diversity among local and exotic accessions. Whereas, 3 SSR primers viz., CaSTMS2, Ca- STMS15 and CaSTMS21 scored the genetic variability up to 55% by cluster analysis through UPGMA percent disagreement. The primers, TA72 and TA130 were linked with yield related traits, indicated highest dissimilarity index value (0.69) and notable variation in the identified promising lines. The Morphometric, Biochemical and Molecular markers reported here, are helpful to assess the extent of genetic diversity among Chickpea accessions and can be used to identify the unreported cultivars with desirable quantitative traits for improving Chickpea yield in Pakistan. Based on the study, the accessions 3043 and 3054 have been recommended to the breeders for their future use in multiplication and screening against various diseases.

Effect of Cultivar, Irrigation and Nitrogen Fertilization on Chickpea (<i>Cicer arietinum</i>L.) Productivity

A 2-year field study was conducted in northern Greece to investigate the effect of nitrogen fertilization and irrigation on productivity of three Greek chickpea varieties (“Amorgos” “Serifos”, “Andros”). Chickpea, grown under irrigation regime (30 + 30 mm of water) and fertilized with 50 kg·N·ha-1 before planting and with 40 kg·N·ha-1 at blossom growth stage, produced more total dry biomass and seed yield as compared with that grown under non-irrigated conditions and fertilized with 50 kg·N·ha-1 before planting only. In particular, irrigation and nitrogen fertilization at blossom growth stage increased total dry weight of chickpea by 18.3% and 18.5%, respectively, as compared with that of non-irrigated and fertilized with N before planting. The corresponding increase of seed yield was 30.5% and 20%, respectively. The total dry biomass of “Amorgos” was 10% and 13% greater than that of “Serifos” and “Andros”, while its respective seed yield increase was 5% and 16%. Finally, the quantum yield of photosystem II of chickpea was not affected by irrigation or fertilization. These results indicated that nitrogen fertilization at blossom growth stage combined with irrigation increased seed yield of all chickpea varieties, whereas the same treatments did not have any effect on plant quantum yield of photosystem II.

Morpho-Anatomical Changes in Roots of Chickpea （Cicer arietinum L.） under Drought Stress Condition

Chickpea （Cicer arietinum L.） is a crop cultivated in semi-arid and rainfed areas of Pakistan and it experiences terminal drought stress. In this paper, the morpho-anatomical study regarding roots of chickpea was carried out to investigate the drought adaptation strategies. Twelve cultivars of chickpea were grown in pots under drought stress of 70% and 35% field capacity in addition to control （100%）. Root segments of mature chickpea plants were sectioned in 2 cm from root-shoot junction and used for making transverse sections. The development of sleeve and stele tissues and their proportion were markedly influenced by moisture availability to the root system. Roots length, flesh and dry weight, and number of secondary roots were decreased under field capacity of 70% and 35% as compared to control in 12 cultivars of chickpea. Root epidermis and cortex of CH47/04 consisted of cells with thickened walls and with 2-3 cell layers of sclerenchymatous cells below the epidermis. Increased number of medullary rays and high vascular region was observed in cultivars CH120/04, CH47/04, CH587/05 and CH87/02. Anatomical studies showed that CH47/04 was highly tolerant among 12 cultivars of chickpea, while CH587/05 and CH87/02 behaved moderately under both levels of drought stress （70% and 35% field capacity）.

RADIATION EFFECT ON BIOSYNTHESES OF ASCORBIC ACID AND RIBOFLAVIN IN GERMINATING CHICKPEA

The influence of irradiation on biosyntheses of ascorbic acid and riboflavin in germinating chickpea seeds at ambient (25-35℃) conditions, was investigated. The rate of syntheses of these vitamins significantly increased with increasing germination time upto 120 h depending upon the treatment (P【0.05). Maximum amounts of ascorbic acid, 22.32 and 16.84 mg/100g, were found in the 0.10 kGy sample after 120 h of germination in tap and distilled water respectively. However, a radiation dose of 0.15 kGy resulted in the development of maximum values of riboflavin, 11.40 and 11.38 μ g/g, on germination in tap and distilled water respectively. A significant linear relation (r = 0-954 to 0.997) was observed between the biosyntheses of these vitamins and the germination time upto 120 h of irradiated and unirradiated chickpea seed (P【0.05).

Mineral concentrations of chickpea and lentil cultivars and breeding lines grown in the U.S.Pacific Northwest

Diseases and health complications caused by mineral deficiencies afflict billions of people globally. Developing pulse crops with elevated seed mineral concentrations can contribute to reducing the incidence of these deficiencies. The objectives of this study were to estimate variance components conditioning seed mineral concentrations of chickpea and lentil grown in Washington and Idaho, determine correlations between different mineral concentrations and between mineral concentrations and yield, 100-seed weight, and days to flowering, and compare seed mineral concentrations between chickpeas and lentils grown in adjacent plots. Genotype effects, although significant in chickpea and lentil for all minerals except selenium, tended to be minimal compared to location, year, and their interaction effects. In both chickpeas and lentils high positive correlations were observed between seed concentrations of phosphorus and potassium, phosphorus and zinc, and potassium and zinc. Correlations between mineral concentration and yield, and mineral concentration and days to 50% flowering were similar for chickpeas and lentils across the majority of minerals. These results may reflect similarities between the two crops in physiological processes for mineral uptake and partitioning. Lentils had higher concentrations of iron and zinc than chickpea when the two crops were grown in adjacent plots,whereas chickpeas had higher concentrations of calcium and manganese. Plant genotypes that are more efficient at obtaining minerals from growing environments will be useful as parental materials to develop improved chickpea and lentil cultivars that have good yield potential coupled with high seed mineral concentrations.

Effect of Plant Density on Yield and Yield Components of Chickpea （Cicer arietinum L.） Grown under Environmental Condition of Golestan, Iran

Chickpea is a major food crop and the world＇s third most widely-grown legume after bean （phaseolus vulgaris L.） and pea （pisum sativam L.）. The yield of chickpea can be improved by planting of optimum density of chickpea cultivars. The purpose of this research was to investigate the effect of plant density on yield and some characteristics of chickpea （Cicer arietinum L. var. Philips）. A field experiment was conducted on research farm of, Gonbad High Education Center, Iran. The study was laid out factorial in randomized block design with four replications. The treatments were factorial row spacing （A1=35 cm, A2=45 cm and A3=55 cm） and plant spacing in the row （Plant-within-spacing row） （B1=5 cm, B2=7.5 cm and B3=10 cm）. AIBI=（35×5）=57.14, AIB2= （35×7.5）=38.09, A1B3=（35×10）=28.57, A2B1=（45×7）=44.44, A2B2=（45×7.5）=29.26, A2B3=（45× 10）=22.23, A3B1=（55×5）=36.36, A3B2=（55 ×.5）=24.24, A3B3=（55 × 10）=18.18 plant.m^-2. At the harvest time, 5 plants of each plot were selected randomly and used for determination of pods per plant, unfilled pod per plant, height and number of branches. Result showed the yield was not affected by density. The maximum yield obtained in row spacing 55 cm and plant spacing in the row 7.5 cm which was equal 1802.87 kg·ha^-1. Row spacing had statistically significant effect on the pod number per plant, branches number per plant and plant height.

Occurrence and Distribution of Chickpea Diseases in Central and Southern Parts of India

A survey was conducted in 2010-2011 rabi cropping season to obtain information on the distribution and incidence of chickpea diseases in respect to soil type, cultivar used, seed treatment in central and southern parts of India. Local cultivars predominated in most farmers’ fields (25% - 48%). 63% of the farmers were practices seed treatment with fungicide. Dry root rot and collar rot diseases were found at all of the sites and incidence ranged from 8.9% - 10.3% and 7.1% - 10.5% respectively irrespective of cultivar type and locations. Incidence of wilt and black root rot disease ranged from 9.7% - 13.8% and 6.6% - 7.4% respectively. Black root rot disease was found in Andhra Pradesh and Karnataka states only. The result indicated that dry root rot and collar rot is currently highly distributed in all surveyed chickpea growing areas of central and southern parts of India. Therefore, possible management options are vital to alleviate the problem.

Unravelling Effects of Temperature and Soil Moisture Stress Response on Development of Dry Root Rot [<i>Rhizoctonia bataticola</i>(Taub.)] Butler in Chickpea

Erratic rainfalls and rise in temperature have become more frequent under the changing scenario of climate particularly in semiarid tropics. As a consequence of it, a drastic shift of chickpea diseases have been recorded throughout the major chickpea growing regions in India and elsewhere. Dry root rot (DRR) caused by Rhizoctonia bataticola (Taub.) Butler [Pycnidial stage: Macrophomina phaseolina (Tassi) Goid] was found as a potentially emerging constraint to chickpea production than wilt (Fusarium oxysporum f. sp. ciceris). Increasing incidence of DRR indicate strong influence of climate change variables such as temperature and moisture on the development of disease. The present study therefore was conducted to quantify the role of temperature and soil moisture associated with infection, colonization and development of DRR under controlled environment. The DRR incidence was significantly affected by high temperature and soil moisture deficit. Out of five temperature regimes (15?C, 20?C, 25?C, 30?C and 35?C) and four moisture levels (40%, 60%, 80% and 100%), a combination of high temperature (35?C) and soil moisture content (60%) predisposes chickpea to DRR. The study clearly demonstrates that high temperature coupled with soil moisture deficit is the climate change variables predisposing chickpea to R. bataticola infection, colonization and development.

Acceptability Properties and Antioxidant Pontential of Desi Chickpea (<i>Cicer arietinum</i>L.) Cultivars

The acceptability properties, total phenolic content, and antioxidant potential of four (red, green, brown, cream) desi chickpea (Cicer arietinum L.) cultivars from the World Germplasm Bank were evaluated. Protein content of the grains varied from 20.12% to 28.85% (dw), highest for Cream ICC3421. The water absorption capacity (WAC) and cooking time (CT) of the whole grains ranged from 97.7 to 117.5 g water/100 g seeds (ww) and from 109.5 to 193.5 min, respectively;the highest WAC and lowest CT corresponded to Cream ICC3421 cultivar. The total phenolic content (TPC) and total hydrophilic antioxidant activity (AoxA) [oxygen radical absorbance capacity (ORAC) value] of desi chickpea cultivars varied from 746 to 1.286 μg gallic acid equivalents (GAE)/g sample (dw) and from 43.9 to 53.9 μmol Trolox equivalents (TE)/g sample (dw);Brown ICC3512 showed the highest ORAC value. Chickpea cultivars could contribute significantly to the management and/or prevention of degenerative diseases associated with free radical damage.

Nature of Gene Action and Maternal Effects for Pod Borer, <i>Helicoverpa armigera</i>Resistance and Grain Yield in Chickpea, <i>Cicer arietinum</i>

Information on mechanisms and inheritance of resistance is critical to plan an effective strategy to breed for resistance to insect pests. Therefore, we evaluated a diverse array of chickpea genotypes (eight desi and one kabuli) with varying levels of resistance to the pod borer, Helicoverpa armigera to gain an understanding of the nature of gene action and possible maternal effects. The test genotypes were crossed in all possible combinations for a full diallel. The 72 F1s (36 direct and 36 reciprocal crosses) along with the parents were evaluated for resistance to H. armigera under field conditions, and for antibiosis mechanism of resistance (larval survival and larval weight gain) by using detached leaf assay under laboratory conditions, and grain yield under un-protected conditions in the field. Additive gene action governed the inheritance of resistance to H. armigera, while non-additive type of gene action was predominant for inheritance of antibiosis component of resistance (larval survival and larval weight) and grain yield. Greater magnitude of σ2 A(17.39 and 1.42) than σ2 D (3.93 and 1.21) indicated the preponderance of σ2 Ain inheritance of resistance to pod borer, H. armigera under laboratory and field conditions, respectively. There were no maternal effects for inheritance of resistance to pod borer and grain yield. Lines with significant gca effects for pod borer damage and grain yield were identified for further use in the resistance breeding program. The implications of the inheritance pattern of pod borer resistance and grain yield are discussed in the context of strategies to enhance pod borer resistance and grain yield in chickpea.

Disease Development Caused by Ascochyta rabiei on Chickpea Detached-Leaves in Petri Dishes

A study using detached-leaves aimed to improve selection method. The improving method was done by scoring both disease symptom and lesion size. The research was begun by selecting agar concentration and dose of conidia that could distinguish response of very susceptible or resistant chickpea genotype. The result was used to determine disease severity (DS) and disease incident (DI) of eight genotypes that were previously tested in the field. Results of the tested agar concentration and dose of conidia showed that 1.5% and 2% agar were good to determine susceptible or resistant genotype;while 1 × 105 or 5 × 104 conidia dose was suitable for inoculation. The formula of DS (no. of leaflets in category × category value/Total no. of leaflets ×10) × 100, and DI (no. leaflets with pycnidial lesions + no. leaflets with necrotic lesions)/Total no. of leaflets × 100 successfully measured genotype response. The lesions development on detached leaves of the susceptible cultivar (Lasseter) began as circular, pale-colored areas, extending to the area covered by the drop of inoculum, then became light brown and finally dark brown. However, the response of resistant line (FLIP508) was restricted in area (and often confined to a tiny speck) surrounded by chlorosis or drying of the tissue.