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Kerala Agricultural University, Thrissur
The history of agricultural education in Kerala can be traced back to the year 1896 when a scheme was evolved in the erstwhile Travancore State to train a few young men in scientific agriculture at the Demonstration Farm, Karamana, Thiruvananthapuram, presently, the Cropping Systems Research Centre under Kerala Agricultural University. Agriculture was introduced as an optional subject in the middle school classes in the State in 1922 when an Agricultural Middle School was started at Aluva, Ernakulam District. The popularity and usefulness of this school led to the starting of similar institutions at Kottarakkara and Konni in 1928 and 1931 respectively.
Agriculture was later introduced as an optional subject for Intermediate Course in 1953. In 1955, the erstwhile Government of Travancore-Cochin started the Agricultural College and Research Institute at Vellayani, Thiruvananthapuram and the College of Veterinary and Animal Sciences at Mannuthy, Thrissur for imparting higher education in agricultural and veterinary sciences, respectively. These institutions were brought under the direct administrative control of the Department of Agriculture and the Department of Animal Husbandry, respectively. With the formation of Kerala State in 1956, these two colleges were affiliated to the University of Kerala. The post-graduate programmes leading to M.Sc. (Ag), M.V.Sc. and Ph.D. degrees were started in 1961, 1962 and 1965 respectively.
On the recommendation of the Second National Education Commission (1964-66) headed by Dr. D.S. Kothari, the then Chairman of the University Grants Commission, one Agricultural University in each State was established. The State Agricultural Universities (SAUs) were established in India as an integral part of the National Agricultural Research System to give the much needed impetus to Agriculture Education and Research in the Country. As a result the Kerala Agricultural University (KAU) was established on 24th February 1971 by virtue of the Act 33 of 1971 and started functioning on 1st February 1972. The Kerala Agricultural University is the 15th in the series of the SAUs.
In accordance with the provisions of KAU Act of 1971, the Agricultural College and Research Institute at Vellayani, and the College of Veterinary and Animal Sciences, Mannuthy, were brought under the Kerala Agricultural University. In addition, twenty one agricultural and animal husbandry research stations were also transferred to the KAU for taking up research and extension programmes on various crops, animals, birds, etc.
During 2011, Kerala Agricultural University was trifurcated into Kerala Veterinary and Animal Sciences University (KVASU), Kerala University of Fisheries and Ocean Studies (KUFOS) and Kerala Agricultural University (KAU).
Now the University has seven colleges (four Agriculture, one Agricultural Engineering, one Forestry, one Co-operation Banking & Management), six RARSs, seven KVKs, 15 Research Stations and 16 Research and Extension Units under the faculties of Agriculture, Agricultural Engineering and Forestry. In addition, one Academy on Climate Change Adaptation and one Institute of Agricultural Technology offering M.Sc. (Integrated) Climate Change Adaptation and Diploma in Agricultural Sciences respectively are also functioning in Kerala Agricultural University.
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ThesisItem Open Access QTL mapping for yield traits in vegetable cowpea(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2017) Ashwin Varghese, V; KAU; Deepu, MathewCowpea [Vigna unguiculata (L.) Walp.] is one of the most cultivated pulse crops in the semi-arid tropics of Asia, Africa, Southern Europe, and other parts of the world. It is used for both vegetable and fodder purpose. In India, kharif crop of vegetable cowpea is cultivated in an estimated area of 0.5 million hectares in states like Kerala, Karnataka, Tamil Nadu and Madhya Pradesh. Studies aimed at increased yield among crops were always challenged by the quantitative nature of traits. These quantitative traits are generally governed by multiple genes present in regions of the genome called quantitative trait loci (QTL). With the advent of molecular markers it is possible to localize the QTL with the help of linked markers, a process now widely known as QTL mapping. QTL mapping depicts the relative positioning of different markers on the chromosomes and their linkage to a specific trait. In cowpea, even though there has been few mapping efforts for traits such as resistance to Thrips tabaci and Frankliniella schultzei, flowering time, pod length and seed weight, an elaborate QTL map for yield and related traits is missing. Hence, the study “QTL mapping for yield traits in vegetable cowpea” was undertaken with the objective of mapping the SSR markers and identifying the quantitative trait loci for yield components in the genome of vegetable cowpea at the Centre for Plant Biotechnology and Molecular Biology (CPBMB), College of Horticulture, during February 2016 to June 2017. F3 plants maintained at CPBMB, derived from the cross of Sharika which is a pole type, long poded, high yielding but anthracnose and cowpea mosaic virus susceptible cultivar with Kanakamony which is a semi-trailing, medium-long poded, low yielding, anthracnose immune and cow pea mosaic virus resistant cultivar, were used to raise the F4 mapping population. Morphological observation for traits pod length, individual pod weight (IPW), pod number, days taken for first flowering (DTFF), total dry pod yield (TDPY), grains per pod, branch number, root length, plant height, plant weight, and response to anthracnose and cowpea mosaic virus diseases were recorded. High quality DNA was isolated from the parents and mapping population using the protocol standardized in this study. One hundred SSR primer pairs reported in cowpea were screened among the parental DNA for polymorphism. Thirty polymorphic primer sets were carried forward to genotype the F4 mapping population. The morphological and genotypic data were used to construct a linkage map using software ICIMapping. Two linkage groups, one having eight SSR markers distributed across 637 cM and another one having five SSR markers distributed across 271 cM were obtained. Two approaches, Single Marker Analysis (SMA) and Inclusive Composite Interval Mapping (ICIM) otherwise called Additive Linkage Mapping were followed for QTL mapping. LOD value threshold of 3.0 was used to determine the significance of QTL and linked markers. Multiple QTL hotspots were observed for different traits under study. An anchored marker, CLM0083 has been identified which was significantly linked to traits individual pod weight and total dry pod yield. The region between 25 cM to 125 cM on linkage group 1 had QTL hotspots harboring genes governing traits DTFF, TDPY, root length, plant length and plant height. This entire region was bracketed by two markers, CLM0244 at 24.25 cM and CLM0177 at 126.86 cM with an anchored marker CLM0008. This marker combination could be potentially used in marker assisted selection for the traits DTFF, TDPY, root length, plant length and plant height. Fine mapping of the QTL for these traits with large number of markers would provide more insights into the genes and hot spots involved in the yield contributing traits in cowpea.ThesisItem Open Access Identification of duplicates in the germplasm of sweet potato (Ipomoea batatas (L.) Lam.) using morphological and molecular markers(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Babitha Babu; KAU; Shirly Raichal, AnilThe study entitled “Identification of duplicates in the germplasm of sweet potato (Ipomoea batatas (L.) Lam.) using morphological and molecular markers” was carried out at the Division of Crop Improvement, ICAR-Central Tuber Crops Research Institute, Sreekariyam, Thiruvananthapuram during 2016-2017. The objective of the study was to identify duplicates in the sweet potato germplasm based on morphological and molecular markers. Identification and elimination of these common redundant materials will enhance the germplasm viability. Fifty accessions were selected for the study. The study was divided into two phases - morphological and molecular analysis. Morphological analysis was performed by using twenty descriptors as provided by IPGRI (CIP et. al., 1991). The recorded data were analyzed statistically by various tools such as PCA and cluster dendrogram. Cluster dendrogram identified three sets of morphological duplicates and the accessions were separated into six principal clusters and two outliers at a Euclidean distance of 1. The PCA analysis revealed predominant vine colour and secondary vine colour, abaxial vein pigmentation and petiole pigmentation as the major factors that contributed to the clustering of the sweet potato accessions. After morphological analysis, molecular analysis was performed. The genomic DNA was isolated using CTAB method which gave good quality DNA. 11 ISSR primers were used for screening of fifty accessions. After the final PCR using selected primers, the product was resolved in 2% agarose and polymorphic bands were obtained. All the primers showed 100% polymorphism and the number of bands ranged from 9 to 18 with a mean value of 14.7 bands per primer. Using the molecular scoring data, UPGMA clustering was done and the whole fifty accessions were divided mainly into two principal clusters and one outlier. The first principal cluster comprised of 40 accessions which were grouped into many subclusters and there was lot of intraclusteral variation. The second principal cluster consisted of 9 accessions and this principal cluster comprised of two true duplicates which were also found similar in morphological characterization. The outlier was different from all the other accessions and may be due to the peculiar leaf shape which is not seen in other accessions selected in the study. SD-29 was different from all the remaining accessions by a similarity coefficient of 0.61.The similarity between the different accessions ranged between 52-100%. The duplicates S-236 and S-256 were 100% similar. The least similar accessions were SD-39 and S-298 (52%). Thus it can be inferred that a 48% variability or diversity existed within the selected accessions which can be considered as a moderate diversity. The hexaploid nature of the crop, self incompatibility, along with the out crossing nature together might have contributed to the high variation observed among the accessions. Only two duplicates were identified. In future more specific markers may be used for core collection development and to eliminate duplicates.ThesisItem Open Access Association mapping for cassava mosaic disease (cmd) resistance in cassava using SSR marker(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Aparna, T K; KAU; Mohan, CThe present study” Association mapping for Cassava Mosaic Disease (CMD) resistance in cassava using SSR marker” Was conducted to identify the marker which are closely associated with CMD resistance in cassava. 30 CMD resistant and 25 CMD susceptible cassava varieties were analyzed with 2n selected SSR primers. Genomic DNA isolated from the leaves and PCR amplification was done with SSR primers. PCR products were separated by PAGE. Polymorphic bands were used to assign loci for each primer and score as presence (1) or absence (0) of bands. RMEI primer showed double banding pattern in 700bp size for all the resistant accessions. But in case of susceptible accessions only single bands were observed. TASSEL and STRUCTURE software were used ro analyses the data obtained from the PAGE.. Cladogram was constructed using genotypic data. Different clusters were formed. Clusters clearly distinguished CMD resistant and CMD susceptible varieties of cassava. CMD resistant varieties sub divided into three sub clusters. Dissimilarity matrix constructed by the software used for the diversity study of collected cassava accessions. Two markers were associated with CMD resistance was identified by the software. SSR36 are the two markers associated with CMD resistance. Result showed that there are eight sub populations in the cassava population used. Based upon the output obtained from Evanno’s method a bar diagram was constructed. Which shows the distribution of genotypes in different sub populations.ThesisItem Open Access Scientific validation of antiinflammatory, antinociceptive and antioxidant potential of malavirinji (Actinodaphne bourdillonii Gamble)(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Adarsh Prathap; KAU; Suja, S RThe study entitled “Scientific validation of antiinflammatory, antinociceptive and antioxidant potential of malavirinji (Actinodaphne bourdillonii Gamble)” was conducted at the Ethnomedicine and Ethnopharmacology Division of Jawaharlal Nehru Tropical Botanical Garden (JNTBGRI), Palode, Thiruvananthapuram, during the year 2016 to 2017. Objective of the study was to scientifically evaluate the antiinflammatory, antinociceptive, antioxidant potential of leaves of an ethnomedicinal plant Actinodaphne bourdillonii Gamble. Phytochemical examination revealed the presence of various phytoconstituents like alkaloids, flavonoids, saponins, alkaloids, carbohydrates and phenols. In in vitro antioxidant method the ethanolic extracts of leaf showed higher free radical scavenging activity compared to standards with IC50 of DPPH, NO Scavenging Activity, Hydroxyl Free Radical Scavenging Activity, Superoxide radical scavenging activity, Total antioxidant activity and Ferric reducing antioxidant potential (FRAP) assay. Toxicity of the ethanolic extract of leaves of A. bourdillonii were tested by acute toxicity study in mice with four doses 25, 100, 400, 1600 mg/kg body weight. And the mice were cage side observed for fourteen days and no toxic effect were seen in the tested animals. In the detailed in vitro pharmacological studies for antiinflammatory and antinociception were conducted. Antiinflammatory activity was determined by Carrageenan (Acute inflammation) and formalin (sub chronic inflammation) induced paw oedema on hind limb in rats with three different doses 50, 150 and 450 mg/kg. At the dose of EAB 450mg/kg give 83.63% of inhibition in carrageenan induced paw oedema and in the formalin induced paw oedema (sub-acute) study a dose of EAB 150mg/kg gives maximum inhibition of 70.23% and 78.36% of inhibition in the first and seventh day respectively. Antinociception activity was determined by Eddy Hot Plate method and Acetic acid writhing method in mice with three different doses 50, 150 and 450 mg/kg. At the dose of EAB 450mg/kg gives maximum inhibition 87.76% of inhibition in Eddy’s hot plate method and in the acetic acid induce writhing study a dose of EAB 450mg/kg gives maximum inhibition of 71.55% of inhibition.ThesisItem Open Access Molecular characterization and in vitro conservation of taro (Colocasia esculenta (L.)Schott)(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Sreevidya, M R; KAU; Asha Devi, AThe study entitled "Molecular characterization and in vitro conservation of taro (Colocasia esculenta (L.) Schott)" was carried out in the Division of Crop Improvement, ICAR – Central Tuber Crops Research Institute, Sreekariyam, Thiruvananthapuram during the period 2016-17. The main objective of the study was to analyze the genetic diversity of taro (Colocasia esculenta) using SSR markers and to maintain them in slow growth conditions for medium term conservation. 36 taro accessions comprising germplasm from various parts of the country, varieties and breeding lines were collected from the taro germplasm maintained at ICAR – CTCRI for molecular characterization and in vitro studies. Good quality and pure DNA was isolated from the selected 36 taro accessions by modified Sharma, et al. (2008) protocol. The quantity of the DNA obtained ranged from 372ng (Line 11) to 2285 ng/μl (DT 1) and the OD values were also good ranging from 0.050 (C-717) to 0.457 (NEH 21). 10 SSR markers were selected for the study, eight from the Uq series (Mace and Godwin, 2002) and two from the Ce1 series (Noyer et al., 2004). The annealing temperatures of these 10 primers were standardized using gradient PCR and ranged from 60 to 68oC. PCR amplified SSR amplicons were resolved in 6% PAGE and 4% high resolution agarose gel electrophoresis. Since, both methods of electrophoresis gave similar results, 4% agarose gel electrophoresis was adopted for further resolution of amplicons from the rest of the primers as it was less cumbersome. The bands obtained were scored as '0','1' based on the presence (1) or absence (0) of bands for further statistical analysis. The SSR primers were found to be highly polymorphic across the 36 accessions and was explained by parameters like Shannon’s diversity index which ranged from 0.88 to 2.09, average number of alleles per locus which ranged from 1.81 to 2.67 and polymorphism information content which ranged from 0.58 to 0.80. The 36 accessions were found to be diverse which was explained by the heterozygosity value (He) which ranged from 0.65 to 0.82. Jaccard’s coefficient was used in NTSys – PC to generate 2 main clusters, I and II. Cluster II had 4 sub clusters. The clustering was not on the basis of geographical similarities as the NEH series which are having geographical similarity were found in different clusters. Common ancestry was also explained when some breeding lines were found in a single cluster. The maximum similarity was found to be 83% between C-557 and NEH 8; NEH 32 and Line 4 as well as Lines 29 and 33. This explained that all the accessions were divergent and there were no duplicates in the set of 36 accessions studied. The main aim of in vitro conservation of taro was to develop an in vitro active genebank of taro and to augment the existing IVAG of tuber crops at ICAR – CTCRI. Apart from conservation, a multiple shoot induction step was incorporated so that rare genotypes present in germplasm or mutants, if present, could be multiplied first before being conserved in the slow growth media. The sprouts from the cormels were taken as explants. The explants were sterilized by keeping in bavistin for 25 min, labolene for 10 min and in HgCl2 for 5 min. They were inoculated in basal MS medium for the establishment of contamination free cultures. The shoot multiplication medium was standardized using six sets of media. Two out of these six media were selected. The best media was MS+TDZ (0.1mg/l) which produced an average of 1.96 multiple shoots and the second best was MS+K (1mg/l) which gave on an average, 1.24 multiple shoots. These cultures were further inoculated into slow growth media, full and half strength MS having 2% sucrose and 2% mannitol. Presence of Mannitol though induced slow growth, explants dried after a few days of inoculation. As a result, for slow growth, full and half strength MS having 2% sucrose was used. Of these two media, half strength MS was found to be more effective in achieving slow growth. The cultures were transferred to the IVAG existing in ICAR – CTCRI. With this study, 36 accessions of taro were found to be diverse with no duplicates being identified and these were kept for medium term conservation in slow growth medium. The study would help the breeders in selecting divergent parents for breeding programs and would help in the conservation of these divergent lines in vitro.ThesisItem Open Access Molecular charecterization of taro bacilliform virus (TaBV)(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Aarathy, M B; KAU; Makeshkumar, TTaro is one of the world’s oldest food crop maintained by farmers for millennia and it has continued to spread throughout the world, as an important crop in Asia, Pacific, Africa and the Caribbean. Viral pathogens are one of the important factors threatening production of this vegetatively propogated crop. Taro viruses are poorly characterized, which is a hindrance to the safe movement of germplasm. The overall aim of this study was to detect the Taro bacilliform virus infecting taro and to characterize the virus at molecular level. Serological detection methods were not employed for the detection of TaBV. Molecular analysis with specific primers (TaBV like sequences) identified a new member among the badnavirus, Taro bacilliform CH virus, which was not previously detected in India. During the study 55 samples from different fields of CTCRI are selected to identify the different symptoms of Taro bacilliform virus infection. The symptoms and occurrence of TaBCHV is still not clear eventhough the samples shows different type of infections with infections in non-symptomatic ones also. Molecular analysis with badnavirus specific primers and further confirmation with PNG BadnaF/PNG Badna R reveals the presence of Taro bacilliform CH virus. It reveals that the RT/RNase H region was highly similar for Taro bacilliform and Taro bacilliform CH virus with <80 % similarity. Sequence and phylogenetic analysis confirms the sequences were more closely related to TaBCHV and exhibited 100 % similarity to TaBCHV isolates TaBCHV-1 and TaBCHV-2. From the Seven pair of primers were specifically designed inorder to characterize the whole genome of Taro bacilliform CH virus three pair of primers were used to successfully characterize the virus partially. To identify the phylogenetic relationship of the sequenced samples with that of available accessions, dendrograms were made using MEGA 5.0 software and the tree showed that sequences has variability eventhough lies within the group. It s clear from the study that the genome sizes of TaBCHV isolates, TaBCHV-1 and TaBCHV- 2 (Gen Bank Accession Nos: KP710178 and KP710177) were 7,641 bp, and it was within the badnavirus genomic range. The two islates almost shared about 98% genomic nucleotide identity. With other reported badnaviruses the genome sequence of TaBCHV-1 and TaBCHV-2 showed similarity to RYNV (44.1 %) and Fig Badnavirus (55.8 %)ThesisItem Open Access Molecular characterization of rice genotypes having Variability in heat tolerance(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Neethu, V Mohan; KAU; Beena, RThe study entitled “Molecular characterization of rice genotypes having variability in heat tolerance” was conducted at the Department of Plant Physiology, College of Agriculture, Vellayani, Thiruvananthapuram. Rice (O. sativa) is very sensitive to high temperature especially at the reproductive and grain filling stage which leads to higher spikelet sterility and ultimately yield losses. The present investigation was, therefore, carried out with the objective to study the variation in rice genotypes for heat tolerance using microsatellite markers. In the present study, a set of 50 SSR primers were employed to assess the genetic diversity among the 10 genotypes. Out of 50 markers, 11 markers showed polymorphism, the marker RM6100 was found as a functional marker associated with heat tolerance in rice, and is functional for further crop breeding programmes. A dendrogram was generated with the aim of analyzing the relationships between the 10 genotypes tested. The genetic similarity index ranged from 0.1 to 0.833. The lowest value 0.1was obtained between PTB7 and CR Dhan202 while highest similarity value (0.833) calculated was between the PTB7 and CR Dhan204 genotypes. However, the genetic diversity analysis with SSR markers will contribute to maximize the selection of diverse parents in the future rice breeding program or development of heat tolerant cultivars.ThesisItem Open Access Metabolite Profiling and gene expression analysis for gingerol production in selected somaclones of ginger (zingiber officinale rosc.)(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2017) Sreeja, S; KAU; Shylaja, M RGinger (Zingiber officinale Rosc.) is one of the oldest known spices and is much valued for its medicinal properties. Ginger rhizome is a source of active biological compounds which are responsible for its medicinal properties. Gingerol is the major pungent polyphenol in ginger which has got a wide array of pharmacological properties. Research on somaclonal variation done at Kerala Agricultural University could locate ginger somaclones with high gingerol content and in the course of investigations high somaclonal variation was observed for quality components. Ginger has a very big genome of 23,618 Mbp which is little exploited and reports on the genomic base of gingerol biosynthesis are scanty. The present investigations hence aim at profiling the metabolites in selected ginger somaclones using high throughput analytical platforms and to analyze the gene expression with respect to gingerol production. One released ginger variety from KAU (Athira), two selected ginger somaclones (B3 and 132M) and parent cultivar (Maran) formed the experimental materials for the study. Studies were carried out at Centre for Plant Biotechnology and Molecular Biology, Distributed Information Centre of College of Horticulture and Arjuna Natural Extracts Pvt. Ltd., Aluva during August 2013 to July 2017. The profiling of aroma principles using Gas Chromatography-Mass Spectrometry (GC-MS) and pungency principles using High Performance Liquid Chromatography (HPLC) at various growth stages viz. five months after planting (5MAP), six months after planting (6MAP) and seven months after planting (7MAP), revealed that aroma and pungency principles accumulated in ginger rhizomes at the rhizome formation stage (5MAP). Clone to clone variation was observed in the number and quantity of aroma and pungency principles accumulated in the rhizome. Total gingerol content in somaclone B3 (19.07%) was high when compared to the control cultivar Maran (17.49%) irrespective of the growth stages. Gene expression for Chalcone synthase in selected somaclones done using real time PCR assay showed highest gene expression in somaclone B3 when the control cultivar Maran was set as calibrator. Somaclone B3 recorded 54 per cent increase in Chalcone synthase gene expression over the control cultivar Maran. Suppression subtractive hybridization done to identify differentially expressed genes in somaclone B3 and control cultivar Maran could prepare Expressed Sequence Tag (EST) libraries both for rhizome and leaf. Analysis of EST sequences (25 rhizome ESTs and 19 leaf ESTs) using various bioinformatic tools revealed that there were no differentially expressed genes for gingerol production in rhizome ESTs. But eleven differentially expressed proteins involved in signaling response, protein trafficking, photosynthesis, ATP formation and transposon mediated mutation were observed in rhizome ESTs. The analysis of leaf ESTs showed differential gene expression in somaclone B3 for 3-ketoacyl CoA thiolase (ACAA1) gene which is involved in gingerol biosynthetic pathway. Hence the higher expression of 3-ketoacyl CoA thiolase gene is responsible for the high gingerol content in somaclone B3 as compared to control cultivar Maran. Eighteen other differentially expressed proteins involved in biological processes like transportation of plant secondary metabolites and their intermediates, mobilization of sucrose into pathways involved in metabolism, lipid biosynthesis, transportation of cellular material to microtubules, biogenesis of metabolic pathways in Calvin cycle were observed in leaf ESTs. The differentially expressed gene (ACAA1) can be further validated using northern blotting and quantitative real time PCR by designing specific primers from the ESTs. Expressed sequence tags and corresponding differentially expressed proteins can be used as molecular markers. Post translational modification in differentially expressed proteins can be used to study the mechanism of gingerol production. Forty four sequences deposited at NCBI form the base sequences for further research.ThesisItem Open Access Tagging of phytophthor pod rot disease resistance gene in cocoa (Theobroma cacao L.) using ISSR markers(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2017) Jeughale Kishor, Pundlik; KAU; Minimol, J SCocoa (Theobroma cacao L.) known as ‘Chocolate tree’, is a major cash crop in tropical countries. Cocoa production is seriously affected by pod rot diseases caused by many Phytophthora species. Among these, the pod rot caused by Phytophthora palmivora has been reported in India. Yearly losses to the cocoa growers around the world from Phytophthora diseases were assessed at 30 per cent of the total yield loss. Disease resistance can be scored using a number of morphological and physiological characters. However, the morpho-physiological characters greatly depend upon the environment which ultimately affect the experimental data. Hence, confirmation of transfer of genes by tagging with the help of a strong tool is of utmost importance in crop breeding. Molecular markers such as Inter simple sequence repeats (ISSRs) have already proven to be a good tool to detect and tag the genes of interest and will help to reduce the breeding cycle. In this context, the present study was taken up with an objective to develop a strategy to tag gene(s) for Phytophthora pod rot (PPR) resistance in cocoa using ISSR markers. Morphological characterization of 28 hybrid progenies of SVI 1.26 × PII 12.11 was carried out by recording five pod and bean characters. High variability was observed for characters viz., pod weight, pod length and breadth, wet bean weight per pod and single dry bean weight among the progeny of the same cross. Detached pod inoculation technique was adopted to classify the hybrids into resistant and susceptible ones. The wide variability was also recorded for disease reaction among the progenies. Based on the resistance score, three resistant and three susceptible hybrids were selected from the segregating progeny. The eight accessions were screened with fifty ISSR and 15 SSR primers to observe polymorphism between resistance and susceptible genotypes. Polymorphism was observed in 11 ISSR primers and from these, six primers viz., UBC 810, UBC 826, UBC 827, UBC 857, Oligo ISSR 04 and Oligo ISSR 08 were eluted and cloned. Plasmid DNA was isolated from clones and sequenced. Though various SSR primer sets screened were found to yield polymorphism, none of them was successful to give a clear distinction among the resistant and susceptible hybrids. This may be due to the fact that, Quantitative trait loci (QTLs) associated with these reported SSR primers may be absent in the genotypes considered for the study. BLASTn analysis specific to plants was done for all six sequences. Upon analysis, Oligo ISSR 04561 had shown 98 per cent identity with Predicted: T. cacao histidine-containing phosphotransfer protein 1 (HPt). HPts play an important role in propagating cytokinin signal transduction. Cytokinins are instrumental in mediating disease resistance by generating a green island around the infection zones, exhibiting delayed leaf senescence and upregulating the expression of the pathogenesis related (PR) gene/s. In addition to this, the auxin-cytokinin antagonism that occurs as part of a complex hormonal interplay, exerts a critical influence on the core SA-JA/ET plant immunity pathways. The BLASTn analysis of marker UBC 810877 resulted in 99 per cent sequence identity with Predicted: T. cacao phospholipid: diacylglycerol acyltransferase (PDAT) 1 mRNA. This protein regulates the synthesis of triacylglycerol, which is a building component of oils in the plant. Accumulation of oil content in plant cells could impart resistance against the pathogen. UBC 827571 had shown 73 per cent sequence identity with T. cacao clone TCC_BA049P20 complete sequence and it is reported to be QTL rich region associated with different traits of T. cacao. Moreover, ISSR markers UBC 810877, UBC 826535 and UBC 857839 are located on chromosome nine, six and four respectively as inferred from NCBI Genome Data Viewer tool through BLASTn annotations. These markers are found to be located in PPR resistance regions rich in defense associated genes. Further validation and exploitation of polymorphic amplicons or markers in response to PPR would be required. The linkage of Oligo ISSR 04561 and UBC 810877 with HPts and PDAT correspondingly have to be validated to elucidate the association and role of cytokinin and triacylglycerol with PPR disease resistance. If validated, UBC 810877, UBC 826535 and UBC 857839 and Oligo ISSR 04561 could be employed as a marker in PPR resistance breeding programmes in cocoa.
