Loading...
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.
Browse
180 results
Search Results
ThesisItem Open Access Cryopreservation of chethikoduveli (Plumbago rosea L.) and assessment of genetic fidelity of regenerated plantlets using molecular markers(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2014) Anand, Vishnu Prakash; KAUInvestigations on “Cryopreservation of Chethikoduveli (Plumbago rosea L.) and assessment of genetic fidelity of regenerated plantlets using molecular markers” were carried out at the Department of Plant Biotechnology, College of Agriculture, Vellayani during 2011-2013. Plumbago rosea var. Agni plants were collected from AMPRS, Odakkali, Ernakulam and maintained at the Department of Plant Biotechnology, College of Agriculture, Vellayani as source of explant during the course of the study. The objectives of the present study was to standardise cryopreservation protocol by encapsulation dehydration technique for long term conservation of P. rosea and genetic fidelity assessment of plantlets recovered and regenerated from cryostorage using molecular markers. The project was carried out in two phases viz., in vitro regeneration and in vitro conservation by cryopreservation of P. rosea. In vitro regeneration protocol was optimised for P. rosea var. Agni. Various steps of in vitro regeneration viz., surface sterilization, axillary shoot proliferation, in vitro rooting and acclimatization and planting out has been standardised. For surface sterilizing, single nodal explants (3-4 cm long) were subjected to fungicide treatment with 0.1 per cent carbendazim 50 per cent W. P. (for 30 min) followed by aseptic sterilisation dip with absolute alcohol. Further, the explants were surface sterilised with 0.2 per cent mercuric chloride (for 5 min) which gave 100 per cent survival without any contamination. Enhanced release of axillary buds from single nodal explants, with maximum shoot proliferation (5.28 shoots/culture) was obtained in the medium, MS + BA 1.5 mg l-1 + IAA 1.0 mg l-1. The best response (10.67 roots/culture) of in vitro rooting of plantlets was obtained in the medium, MS + NAA 1.0 mg l-1. In vitro rooted plants gave a maximum survival rate of 76 per cent and 72 per cent, when planted out in potting media consisting of red soil and coir pith (3:1) and red soil and coir pith (2:1) supplemented with VAM respectively at 25 per cent shade. In cryopreservation studies, preconditioning treatment (sucrose 0.5 M for 7 days) recorded maximum shoot proliferation (2.67 shoots/culture) when nodal segments with single axillary bud were cultured on MS + BA 1.5 mg l-1 + IAA 1.0 mg l-1 medium. Among different encapsulation treatments, maximum shoot proliferation of (2.31 shoots/culture) was obtained in beads formed with sodium alginate 2.5 per cent and calcium chloride 100 mM, when cultured on the medium, MS + BA 1.5 mg l-1 + IAA 1.0 mg l-1. Pre-culture medium supplemented with sucrose 0.5 M for 3days gave maximum shoot proliferation (3.44 shoots/culture) when cultured on the medium, MS + BA 1.5 mg l-1 + IAA 1.0 mg l-1. A desiccation duration of 5 h at 18.13 per cent moisture level was found to be most effective giving 66.67 per cent survival and 62.50 per cent regeneration on thawing and culturing on the recovery medium MS + BA 1.5 mg l-1 + IAA 1.0 mg l-1. The beads when stored in liquid nitrogen for different duration and cultured on recovery medium did not show any significant variation with respect to survival per cent. RAPD markers were tried to study the genetic fidelity of the regenerated plantlets from encapsulated and cryopreserved axillary buds. Six primers were screened and RAPD banding patterns of the cryoregenerated plantlets and control plants were compared. Polymorphism was not found with any of the primers tested. RAPD profiles of cryoregenerated plantlets were identical to those of the control. The in vitro regeneration protocol optimized included surface sterilization of single node cuttings with 0.2 per cent HgCl2 for 5 min, axillary shoot proliferation in MS medium supplemented with BA 1.5 mg l-1 and IAA 1.0 mg l-1, in vitro rooting in MS medium supplemented with NAA 1.0 mg l-1 and planting out in potting medium, red soil and coir pith (3:1). The protocol for encapsulation dehydration technique of cryopreservation was standardised for the axillary buds of P. rosea with preconditioning in semi solid MS medium supplemented with sucrose 0.5 M for 7 days, encapsulation using sodium alginate 2.5 per cent and calcium chloride 100 mM followed by pre-culture in liquid MS supplemented with sucrose 0.5 M for 3 days and 5 h dehydration (MC 18.13 %), rapid freezing in LN for at least 2 h and recovery in the medium MS + BA 1.5 mg l-1 + IAA 1.0 mg l-1. The cryopreservation protocol using encapsulation-dehydration technique standardised could be utilised for long-term conservation of P. rosea.ThesisItem Open Access Somatic embryogenesis from immature inflorescence of coconut (Cocos nucifera L.)(Department of Plant Biotechnology, College of Horticulture, Vellanikkara, 2019) Radhika, R; KAU; Sujatha, RCoconut palm (Cocos nucifera L.), the only species of genus Cocos, is a major crop plant which is propagated exclusively by seeds presently. Coconut is highly heterozygous, generally cross-pollinated and long lived. So the offspring differ greatly due to segregation, which can be assessed only after a very long juvenile phase. Breeding for crop improvement is thus a difficult and long-term process. So within a short period, in- vitro propagation, employing tissue culture techniques, offers a means of cloning improved planting material. So the objective of the study was to assess the in vitro response of immature inflorescence of coconut explants for induction and germination of somatic embryos by supplementing the medium with by-product of neera processing and neera. The media for culture initiation, subculturing and embryo germination were standardised in earlier experiments (Siny, 2006; Sujatha, unpublished data, 2011). In the present experiments, standardised sub-culturing medium with addition of different supplements were tested. Embryogenic calloids and somatic embryos were induced in standardized CM2 media (Y3 basal media containing a combination of 0.5 mgL-1 picloram, 1 mgL-1 NAA and 0.1 mgL-1 TDZ and 300-600 μM 2,4-D) from the immature inflorescence explant. Among the different stage and region of immature inflorescence tested in standardised CM2 medium, tip region of inflorescence (85.71%) with length 1-5 cm was found to be the best for calloid induction with low browning (4.76%). The adverse effect of exudation of polyphenols was reduced by inoculating youngest inflorescence with reduced thickness. Embryogenic calloids were sub- cultured into 10 media combinations. With four stages of explant (based on length), each stage with two types (tip or base) there were eighty combinations. The suitability of the by-product from neera processing as well as neera (CIS- Coconut Inflorescence Sap) as supplements for CM2 medium to culture the immature inflorescence of coconut was assessed and found that CM2 medium with 0.3 per cent activated charcoal, 4 per cent sucrose and 2 per cent supplement and also CM2 medium with 0.3 per cent activated charcoal, 4 per cent sucrose and 10 mlL-1 CIS was the best medium for obtaining cultures with calloid proliferation (92.75 % & 90.17 %) with minimal phenolic interference (11.58 % & 22.7%). Calloid proliferation was successfully achieved from all the media combinations. Chemical properties of neera supplement used in the culture mediumThesisItem Open Access DNA barcoding of Spider mites (Prostigmata:Tetranychidae) associated with Ornamental Plants(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2019) Jayalakshmi, Prakash; KAU; Haseena, BhaskarTotal area under floriculture in India is estimated to be 0.31 million hectares with a production of 2.08 million tonnes loose flowers and 0.82 million tonnes cut flowers in 2018-19. In Kerala, Thrissur district is considered as the centre of floriculture nursery business. Majority of nurseries in the area do not maintain their own sources of mother plants and hence import planting materials from other countries. Trade of commercial ornamentals has been recognized worldwide as an important invasion pathway for non-native pests, especially insects and mites. Though mites are reported as major pests of commercial ornamental plants from different parts of India, no systematic study has been conducted so far to document the diversity of mites associated with ornamental plants in Kerala. Considering this, the present study, ‘DNA barcoding of spider mites (Prostigmata: Tetranychidae) associated with ornamental plants’ was undertaken with the objectives to generate DNA barcodes for different species of spider mites infesting commercial ornamental plants of central Kerala and to find the genetic variability among them. The study included collection and culturing of spider mites, morphology based identification and molecular characterization of selected accessions. Purposive sampling surveys were carried out in commercial ornamental nurseries and homestead gardens of Thrissur and Ernakulam districts, covering 12 ornamental plants. Mite infested leaf samples were collected, brought to the laboratory and maintained separately as isoline cultures by assigning unique accession numbers. Morphological characterization of 26 isoline cultures revealed the occurrence of six species of spider mites under two genera viz., Tetranychus and Oligonychus. The genus Tetranychus was more diverse with five species viz., Tetranychus truncatus, T. urticae, T. okinawanus, T. neocaledonicus and T. marianae. The genus Oligonychus was represented by only one species, Oligonychus biharensis Hirst. Rose recorded the highest diversity of spider mites with five species. The mite species, T. okinawanus recorded wider host range with eight host plants viz., Adenium, rose, Gerbera, chrysanthemum, orchid, cairo morning glory, marigold and balsam. All the host plants except Adenium are new host records of T. okinawanus from India. The study recorded three new host plants for T. truncatus from India viz., rose, cock’s comb and marigold. In this study, T. marianae was recorded for the first time from Kerala. For molecular characterization, DNA was isolated and COI locus of 868 bp length was amplified using universal primer, specific to COI. Polymerase chain reaction (PCR) products of 12 accessions representing five species were sequenced and in-silico analysis was carried out. Homology analysis of sequences of 11 accessions showed 90-99 per cent similarity with sequences in NCBI database, which were in consensus with morphological identification. The sequence of the accession, Ros1vk (T. marianae) did not show significant similarity with any of the sequences in the NCBI database. In this study, COI sequence of T. marianae was submitted for the first time in GenBank. Barcode gaps among the species were examined by aligning the COI sequences using Clustal Omega tool and species-specific barcodes were identified at different nucleotide positions. Pairwise distance analysis of the sequences showed intraspecific divergence ranging from 0.00 to 0.036 and interspecific divergence ranging from 0.070-0.217. Phylogenetic analysis revealed the monophyly of T. truncatus and T. urticae and the close relationship of T. marianae with T. okinawanus. The study has shown that rose harbours many species of spider mites, indicating the need for imposing strict quarantine regulations for movement of planting materials of rose to avoid entry and invasion of mites into newer areas. The potential of T. okinawanus and T. truncatus to turn invasive in Kerala’s ecosystems is also brought out. The study establishes the reliability of COI locus as a marker for species discrimination in spider mites.ThesisItem Open Access Inheritance of molecular markers linked to vascular streak dieback disease resistance in hybrid progenies of cocoa (theobroma cacao L.)(Department of Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2019) Midhuna, M R; KAU; Minimol, J STheobroma cacao L. (also known as the chocolate tree) is a major cash crop and the costliest beverage crop. Andhra Pradesh is the leading cocoa producing state in India but Tamil Nadu ranks first with an area of 26,969 ha. Vascular Streak Dieback (VSD) caused by the fungus Ceratobasidium theobromae is a serious disease in cocoa. Since it is a vascular pathogen, chemicals have little effect on disease control. The most tenable and economic technique to tackle this disease is by evolving resistant materials. Kerala Agricultural University had initiated VSD resistant breeding since 1995. Seedlings from hybridization, exhibiting high levels of resistance were selected and field established. Nineteen hybrids, exhibiting resistance to VSD (after screening for a period of thirteen years), were selected for the present study. The progeny obtained from these hybrids by crossing it among themselves were used as plant materials for the study. Two thousand two hundred and thirty seven flowers were pollinated and seven pods were obtained. About two hundred and sixty nine seedlings were grown from the seven hybridized pods in which nursery screening for disease resistance was done. Inoculum was dispensed by keeping already infected seedlings around the experimental materials. High humidity was ensured by providing over head sprinkler system. Visual screening recorded one hundred and eighty seedlings as disease resistant, fourteen seedlings as partially resistant and seventy five seedlings as disease susceptible. Three ISSR markers (UBC 811, UBC 815 and UBC 857) and one SSR marker (mTcCIR42) linked with VSD resistance gene, identified and validated from the previous studies were used for screening of the one hundred and twenty seedlings out of which one hundred and six were resistant and fourteen were partially resistant. The polymorphic band of 950 bp, which was found to be linked with the gene conferring VSD resistance was recorded in seventy seven resistant seedlings and three partially resistant seedlings, when screened with the primer UBC 811. When screened with the primer UBC 815 (750 bp) and UBC 857 (450 bp), the polymorphic marker band which was found to be linked with VSD resistant gene from the previous studies, was present in only twenty five resistant and one partially resistant seedling and twenty one resistant and one partially resistant seedling respectively. When screened with SSR marker, the 200 bp marker band, which was tagged with the VSD resistant gene was detected in fourty six resistant and six partially resistant seedlings. The ISSR marker UBC 811 and SSR marker mTcCIR42 were found to be having comparatively good percentage of inheritance among the segregating progeny screened with a mean inheritance percentage of 71.70 per cent and 48.78 per cent respectively. Flanking sequences of the ISSR markers UBC 811, UBC 857 and SSR marker mTcCIR42 were extracted from the whole genome database of cocoa. The ORFs from the flanking sequences of UBC 811 were identified to be uncharacterized proteins by using BLASTp tool. One ORF from the upstream sequence of the UBC 857 had shown identity with beta tubulin chain. Analyzing the distance between the marker and the flanking region, it was deduced that UBC 857 is a part of beta tubulin gene. Two ORFs were identified from both the upstream and downstream flanking sequences of the SSR marker. Using BLASTp tool, it was analyzed that both the ORFs showed more than 97 per cent identity to beta tubulin gene. Analysing the spacing between the marker and the flanking sequences, it was deduced that both the ORFs are part of the same gene and the SSR marker mTcCIR42 lies within the beta tubulin gene. Tubulin beta chain belongs to the microtubular component of cytoskeletal elements which provides resistance by not allowing the fungi to penetrate the outer epidermal wall of the plants, hence protecting the plants from infection. The ISSR marker UBC 857 and the SSR marker mTcCIR42 are linked to the beta tubulin gene, which provides VSD resistance by giving resistance against penetration of the plant cell by the fungus. Nineteen seedlings were identified to be having three or more markers expressed. They can be planted in the field and can be further evaluated for yield and yield contributing characters. Sequence of the beta tubulin gene can be used for primer designing, which can be used for confirmation by screening in resistant genotypes of cocoa.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 Development of an in vitro regeneration system and validation of genetic stability in phalaenopsis hybrid winter spot with molecular marker(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2016) Asha Amal, Raj; KAU; Lissamma, JosephPhalaenopsis “Moth Orchids” are among the most beautiful flowers in the world. This genus has economic value for pot plant and cut flower production and is distributed throughout Southeast Asia. Most popular method of propagation for orchid is through in vitro propagation, as it produces large number of clones in relatively short duration. Despite its potential to produce numerous plants from a single leaf segment, it is liable to unpredictable mutations or somaclonal variation during the process of multiplication. Variation can arise due to many reasons such as type of media, plant growth regulators and its concentration, type of explants and number of subculture cycles. The percentage of the variation can range from 0-100% depending on varieties with an average of 10% among Phalaenopsis (Tokuhara and Mii, 1993). So the present investigation on “Development of an in vitro regeneration system and validation of genetic stability in Phalaenopsis hybrid Winter Spot with molecular marker” was taken up at the Center for Plant Biotechology and Molecular Biology, College of Horticulture from 2013-2016. Flowering mother plants of Phalaenopsis hybrid Winter Spot were used as explant source. Among the explants namely inflorescence node, transverse thin cell layer of leaf and root segments used for tissue culture study in this orchid, inflorescence node was the best with respect to culture response. The best surface sterilization treatment for leaf explants identified was treatment 0.1% bavistin + prill 2 drops (30 min) and 0.1 per cent HgCl 2 ( 8 min) which give maximum per cent of culture survival and minimum contamination rate. The best surface sterilization treatment for inflorescence node identified was treatment with 0.1% bavistin + 2 drops prill (30 min) , one minute dip in 70 per cent ethanol and 0.1% HgCl 2 (7 min).From different basal media (full MS and 1⁄2 MS) tried, response was observed only in the medium of Full MS for inflorescence node. Among the different growth regulators tried, MS medium supplemented with BA and TDZ was found to give good shoot regeneration from inflorescence node explants. MS +2mgl -1 TDZ recorded highest percentage (80%) of culture establishment, followed by MS + 4.5 mgl -1 of BA (55%) per cent of sprouting. Among the explants tried, only inflorescence node responded with sprouting. Root segment remained as such without any change, whereas leaf explants remained green up to 2 weeks, thereafter started drying in all the growth regulators combination. For induction of multiple shoot, MS medium supplemented with 4.5 mgl -1 BA resulted in the highest average number of multiple shoot (4.15). Elongation and rooting was observed in MS medium supplemented with BA 4.5mgl-1 +IAA 1mgl -1 with 80 % rooting. Root initials were observed 50 days after inoculation. The potting media, charcoal, brick pieces and sphagnum moss in the ratio of 1:1:1 was found ideal for hardening of Phalaenopsis hybrid winter spot with 100% survival. Genetic stability studies using RAPD marker were carried out with the mother plants along with three regenerants each. Six primers were selected based on DNA amplification pattern. In RAPD assay, M1 mother plant recorded the highest average polymorphism of 19.7% and M3 mother plant recorded the least average polymorphism of 8.18%. Using NTSYS software, the similarity coefficients for first, second and third plant between M1 mother plant, M2 mother plant and M3 mother plant and corresponding regenerants were 0.91, 0.92 and 0.93 respectively. In fourth plant, the similarity coefficient exhibited 100% similarity between mother plant, the first clone C1 and third clone C3. The established micropropagation protocol can be used with suitable modification for large scale production of other Phalaenopsis varieties.ThesisItem Open Access Evaluation of miRNA prediction tools and in silico analysis of micro and long non coding RNAs in sweet potato(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2018) Aswathy, M B; KAU; Sreekumar, JThe study entitled “Evaluation of miRNA prediction tools and in silico analysis of micro and long non coding RNAs in sweet potato (Ipomoea batatas L.)” was conducted at the ICAR-CTCRI, Sreekariyam. The objectives of the study is to compare different miRNA and target prediction tools and in silico analysis of the miRNAs and lncRNAs in sweet potato. The plant miRNA identification tools: NOVOMIR and miRPlant and miRNA-target prediction tools: psRNATarget and miRanda were compared. NOVOMIR and psRNATarget were found to be a better tool in miRNA identification and target prediction. MicroRNAs (miRNA) are 18-22nt small, endogenous non coding RNA that has prominent role in many biological processes. In the present study, we report the computational prediction of miRNAs and targets from expressed sequence tags (ESTs) of sweet potato. We predicted 13 novel potential miRNAs and 81 potential target genes and functionally characterized by BLASTX and BLAST2GO. The predicted target genes were credited with their role in signalling cascades, metabolism, and defence and stress responses. Another candidate that has more importance in the genome regulation is lncRNAs. lncRNAs are greater than 200 nucleotide length ncRNA candidate that holds functions at RNA level itself. RNAplonc is a plant long non coding RNA identification tool which uses 16 feature selection methods to predict long non coding RNA molecules. The present study which predicts 9215 lncRNAs and 8665 protein coding genes by RNAplonc in sweet potato for the first time using available ESTs sequences. Since there is a lack of lncRNA functional annotation tool, the functional analysis of predicted lncRNAs is quiet difficult. From the predicted miRNAs and lncRNAs two miRNAs and two lncRNAs were randomly selected for experimental validation by real time quantitative PCR using three different sweet potato varieties Sree Kanaka, ST13 and Khanjakad available at ICAR-CTCRI and compared the target gene’s expression in each variety. Validation results prove that both the miRNAs and lncRNAs shows their importance in crop improvement.ThesisItem Open Access Characterisation of Pathogenesis related proteins for anthracnose resistance in vegetable cowpea, Vigna spp.(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2013) Agatha Shiny, A; KAU; Deepu MathewCowpea (Family: Fabaceae) is an important pulse cum vegetable crop of suitable for the tropical and sub-tropical regions of the world. The grain type cowpeas better tolerates the biotic and abiotic stresses against the vegetable types. Under humid conditions, vegetable types, especially the pole types are susceptible to many diseases and among them, anthracnose caused by Colletotrichum lindemuthianum (Sacc. & Magn.) Br. and Cav. is very severe. In Kerala, complete yield loss in vegetable cowpea is reported due to anthracnose during monsoons. The study entitled “Characterization of pathogenesis related proteins for anthracnose resistance in vegetable cowpea, Vigna spp.” was carried out with objective to develop the protein profiles of resistant and susceptible bush and pole genotypes through SDS-PAGE analysis at different time intervals of infection and to characterize the differentially expressed proteins by MALDI-TOF followed by in-silico analyses. Two bush type varieties Pusa Komal and Kanakamony, the former reported to be highly susceptible and the latter immune to anthracnose and two pole type varieties Lola and Arimbra Local, of which the former susceptible and the latter resistant were used in the study. Pure culture of the pathogenic fungus was developed and maintained on selective medium (Neopeptone-Glucose-Agar) at the Dept. of Plant Pathology. The identity of Colletotrichum lindemuthianum has been established from the spore characteristics observed under phase contrast microscope and the pathogenicity was confirmed through artificial inoculation under controlled conditions. The pot culture experiment was conducted with 50 pots per variety. Artificial inoculation of pathogenic fungus was done and the leaf samples were collected at 0, 6, 12, 18, 24, 48, 72, 96, 120, 144,168 and 192 hours after artificial inoculation. The total protein was extracted using Tris-HCl buffer (pH-7.5), quantified using spectrophotometer and analyzed by SDS-PAGE method. The defense enzymes like peroxidase (PO), polyphenol oxidase (PPO) and phenylalanine ammonia-lyase (PAL) were assayed. By artificial inoculation, disease responses for anthracnose were confirmed to be highly susceptible in Pusa Komal and Lola; highly resistant in Arimbra Local and immune in Kanakamony. Protein expression was found to be higher from the initial hours in resistant varieties whereas in susceptible varieties, the expression was reduced immediately after infection then peaked at 18hr and gradually decreased later on. Two prominent and differentially expressed protein bands at 56 kD and 14 kD were sequenced in MALDI-TOF to obtain the peptide mass fingerprint. Through in-silico analyses using Mascot server software, they were identified to be the large and small subunits of the chloroplastic enzyme RuBisCo. Thus the capability of a variety to maintain high levels of RuBisCo was found to be the deciding factor for anthracnose disease resistance. Further, protein profiles developed after purification of proteins by dialysis have clearly identified the differentially expressed band at 29 kD in the resistant varieties which is in the size range of already reported PR proteins. PO and PAL activities were proportionate to the resistance behavior, with the peak values at 18 and 24 hr after inoculation. With the results of this study, these defense enzymes are recommended as biochemical markers for identifying the resistance in the accessions. Capability to maintain higher levels of RuBisCo, PO and PAL enzymes is the characteristic of anthracnose resistant vegetable cowpeas and the future breeding programmes could be oriented in this directionThesisItem Open Access DNA barcoding in genera benincasa and praecitrullus(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture,Vellanikkara, 2019) Priya Sonkamble, KAU; Narayanankutty, CBenincasa is a monotypic genus with only one cultivated species hispida. In India, Benincasa exhibits rich diversity. There is substantial variation in vegetative traits and fruitcharacters and it is difficult to distinguish ashgourd genotypes based on their external morphology alone. In spite of having enormous morphological variability, all members at present are accommodated into asingle species. The taxonomic classification of Praecitrullus is still uncertain. The marker and pollen morphology studieshave revealed that P. fistulosusis more related to B. hispida, than compared to other cucurbit species. DNA barcoding is a novel system designed to provide rapid, accurate and automatable species identification using short, standardized genomic regions as internal species tags. DNA barcoding is based on the characteristic variations on the sequences of identified genomic regions, which can distinguish individuals of a species. Species identification through barcoding is usually achieved by the retrieval of a short DNA sequence from a standard part of the genome (i.e. a specific gene region either from chloroplast, mitochondria or nuclear genome) and identifying the barcode gaps for each species. The barcode sequence from each unknown specimen is then compared with a library of reference barcode sequences derived from individuals of known identity. The study entitled “DNA barcoding in genera Benincasa and Praecitrullus was done at Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, with the objectives to assess genetic diversity and interspecies relationship among Benincasa and Praecitrullus genotypes and to check the species status of Benincasa using barcodes. Twenty-six ashgourd and five tinda accessions were morphologically screened in the field using the standard descriptor and basedaccessions were clustered, based on morphological characters recorded. The ashgourd accessions varied widely for their morphological traits,fruit size varied between0.191 to 12.0 kg. Thefruit shape, varied from club, spindle, cylindrical, oblong to ridged shape. The growth habit varied from long viny to short viny.Serrated, deep serrated and double serrated leaf margin was observed. For stem pubescencevariations recorded were hispid, downy, tomentose, puberulent, pilose,villous and hirsute. The cross compatibility of the ash gourd accessions were also checked to establish the species status. All the accessions of ash gourd were cross compatiblein both the directions, showing that they belong to the same species. Based on the morphological data of ashgourd accessions nine cluster were generated. One representative accession from each cluster and three representative accessions of tinda selected for the molecular studies. Total genomic was isolated and subjected to PCR assay. The matK and ITS2 gave the bands of 950 bp and 799 bp respectively, which were sufficient to identify barcode gaps.The markers were sequenced from 9 genotypes of ashgourd and 3 tinda genotypes.The BLAST analysis had shown that matK and ITS2 both loci is 99 per cent efficient for species discrimination in Benincasa and Praecitrullus. Barcode gap, a position in the sequence at which a particular nucleotide shows characteristic variation in all the members of a particular species, was identified for all the members of Benincasa and Praecitrullus species. Barcode gaps were identified for tinda, in comparison with Benincasa for both the loci. The barcodes in tinda were observed in matK locus at 47, 127 and 232bp. Similarly, for the locus ITS2, the barcode gaps in tinda were identified at 162, 207, 278, 279 and 288 bp. However, within Benincasa, there were no such characteristic barcode gaps, indicating that the null hypothesis of monotypic status of Benincasa is true. Phylogenetic analysis using Clustal Omega showed the variation within Benincasa species and Praecitrullus species.The barcodes developed in this study could be successfully used to solve the genetic relatedness of Benincasa and Praecitrullus species.
