1. KAUTIR (Kerala Agricultural University Theses Information and Retrieval)
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Item Characterization of selected curcuma species germplasm using morphological and molecular markers(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2018) Bimal Thomas; Asha, K ICurcuma L., a perennial rhizomatous herb, is gaining global importance as a source of starch besides its medicinal property and use as a spice. Characterization of germplasm is very essential in crop plants and it is the basis for selection of accessions for use in crop improvement programmes. This research work was an attempt to characterize the fifteen selected accessions in eight species of Curcuma collected from different parts of India and maintained in the field gene bank of ICAR-CTCRI using morphological and molecular markers. Two accessions in each of C. amada, C. angustifolia, C. aromatica, C. decipiens, C. malabarica, C. raktakanta, C. zedoaria and one of C. longa were selected. These 15 accessions were morphologically characterized using 13 qualitative and 15 quantitative traits and a wide variability was observed. Dendrogram based on the morphological characters grouped the genotypes into four clusters. PCC analysis revealed that the accessions of the same species have shown more than 83% similarity except C. angustifolia. C. raktakanta accessions have shown a highest intra-specific similarity of 94%. C. decipiens accessions were found to be the highly variable from the most commonly exploited species C. longa while C. aromatica has shown highest similarity. PCA showed that the characters such as leaf midrib colour, rhizome flesh colour, leaf texture and aroma of rhizome have contributed mostly to the variability. Molecular characterization was done using 10 ISSR and 7 SSR markers. The total percentage polymorphism obtained by ISSR characterization was 94.31 while it was 91.11 percentage in the SSRs. C. angustifolia-1 was found to be highly variable from C. angustifolia-2 suggested the occurrence of intraspecific variability. The intra-specific similarity among C. raktakanta accessions were found to be highest than all other accession pairs. Clustering based on ISSR markers grouped the genotypes into five clusters while SSRs into six clusters. Mantel’s test showed a positive correlation between the morphological and molecular data. The results of the present study indicated that the morphological as well as the molecular tools were found to be very effective in the characterization of germplasm of Curcuma species for the developement of core collections and for further use in the crop improvement programmes.Item 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; 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.Item Characterization of selected accessions of cassava germplasm using morphological and molecular markers(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Anjali sabu, C; Asha, K ICassava (Manihot esculenta Crantz), a perennial shrub, is an important crop in many parts of the tropics.This research work attempts morphological and molecular characterization of 27 cassava germplasm collected from Southern India. In the present study27 accessions of cassava maintained in the field genebank of ICAR- Central Tuber Crops Research Institute, Sreekariyam, Thiruvananthapuram were characterized based on 20 qualitative and 10 quantitative traits including the major yield components. There were no duplicate accessions identified based on morphological classification and it can be maintained as core collection. The genetic diversity on molecular basis was evaluated using 10 SSR primers. Molecular markers are proved to be valuable tools in the characterization and evaluation of genetic diversity within and between species and population. All the SSR primers in the study showed the polymorphism. The SSR primers on an average produced 9 polymorphic alleles with mean observed hetrozygosity and values of Polymorphism Information Content (PIC) 0.8293 and 0.8091 respectively. The value of heterozygosity here ranged from 0.2975 (SSRY 148) to 0.8293 (SSRY 9). The PIC value ranged from 0.2533 (SSRY 148) to 0.8091 (SSRY 9). The hetrozigosity and average PIC content observed in SSRY 9. Clustering based on morphological descriptors and molecular markers was done. In morphological clustering, Cluster-I consists of 7 accessions was further subdivided into two sub clusters I A and I B. Cluster-I A consisted of four accessions while Cluster-I Bof three accessions. The bigger cluster Cluster-II consisting of 20 accessions was found to have two subgroups namely II A with 12 accessions and II B with 8 accessions. Clustering based on SSR marker analysis grouped the genotypes into 2 Clusters. Cluster-I contain 6 accessions was further sub grouped into I A and I B with 3 accessions each. Cluster II with 21 accessions was found to have two subgroups II A with 13accessions and II B with 8 accessions. By comparing the morphological and molecular clusters, In Cluster II of each dendrogram have 6 similar accessions of cassava (TCR-5, TCR-10,TCR-15,TCR- 45,TCR-79,TCR-69,). Clustering and Principal Componant Analysis of the data validated the variation among the cassava accessions. In PCA of morphological characters the percentage variation obtained in PC component 1 (54.22). Mantel’s test proves that there is no correlation between the morphological and molecular data. The present results indicated that the primers selected for the present study will be useful for future genetic variability studies and would provide breeders with a genetic base for selection of diverse parents for crop improvement programmes in cassava.Item Molecular analysis of spike branching observed in black pepper (Piper nigrum L.) type from Idukki(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2009) Vimarsha, H S; Rajmohan, KThe study entitled “Molecular analysis of spike branching observed in black pepper (Piper nigrum L.) type from Idukki” was conducted at the Department of Plant Biotechnology, College of Agriculture, Vellayani, Thiruvananthapuram during 2006-2008. The objective of the study was to analyse the sequence homology for the inflorescence architecture gene (TFL1) and flowering gene (FT) in spike branching black pepper type from Idukki, using polymerase chain reaction technique. Spike branching in black pepper is very rare phenomenon. Profuse spike branching was observed in an uncharacterized black pepper type from Idukki. Single spike had three to four times more berry yield, compared to improved varieties. This type needed to be characterized at morphological and molecular levels to know its parental descendent and possible involvement of gene regulation responsible for spike branching. As an initial approach, molecular analysis was done to find out the presence of TFL1 and FT genes, which have been reported to be involved in inflorescence branching in modal plants. TFL1 primer pair (TFL-F1 and TFL-R1), designed for the fourth exonic sequence of TFL1 gene in Arabidopsis thaliana, could amplify a 700 bp DNA sequence in the spike branching type, as well as in Karimunda and Vellamundi, indicating homology for the gene sequence and possible conserved exonic sequence in black pepper. Presence of sequence homology for the gene TFL1 indicated the possible involvement of TFL1 gene, which had been reported to be associated with inflorescence branching in Arabidopsis thaliana, in the spike branching trait of the black pepper type. This result is significant, as five out of the eight cultivars tested did not give any positive response for the primer pair designed based on TFL1 gene. However, two non-spike branching varieties including Karimunda and Vellamundi have also shown amplification for the TFL1 primer pair. As an initial step towards characterisation of the spike branching type, RAPD analysis of it along with seven other cultivars/varieties was done. This analysis revealed variability, accounting for 83 per cent polymorphism. In the dendrogram, at a similarity index of 0.43 the plants grouped into two big clusters, indicating 67 per cent dissimilarity. All the eight plants under study formed individual clusters at similarity index 0.74, except the spike branching type and Vellamundi. Results of gene specific PCR which yielded single amplicon can be hypothecated for the presence of sequence homology for the gene TFL1 and has conserved fourth exon in black pepper cultivars and varieties. Further transcript level and expression level studies are essential to find the specific role of the TFL1 gene in spike branching black pepper.