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Open Access
MOLECULAR MARKERS BASED GENETIC DIVERSITY ANALYSIS IN FINGER MILLET [Eleusine coracana (L.) Gaertn.] 3689
(JAU,JUNAGADH, 2023-06) DUDHATRA RUTU JAYESHBHAI; Dr. S.B. Bhatt; 2010121018
The present investigation on “MOLECULAR MARKERS BASED GENETIC DIVERSITY ANALYSIS IN FINGER MILLET [Eleusine coracana (L.) Gaertn.]” was conducted at Department of Biotechnology, Junagadh Agricultural University, Junagadh with objectives to analyze different finger millet (25) genotypes for biochemical and molecular diversity using various PCR based molecular markers viz. Random Amplified Polymorphic DNA (RAPD), Inter Simple Sequence Repeats (ISSRs) and Simple Sequence Repeats (SSRs) as well as to find out the phylogenetic relationship among different finger millet genotypes.
Open Access
MOLECULAR CHARACTERIZATION, NUTRITIONAL AND ANTINUTRITIONAL PARAMETERS OF PROMISING GENOTYPES OF BLACK GRAM [Vigna mungo (L.) Hepper] 3571
(JAU,JUNAGADH, 2022-09) PATEL KALPANABAHEN KHEMABHAI; Dr. U. K. Kandoliya; 2010120068
The present investigation on “Molecular Characterization, Nutritional and Antinutritional Parameters of Promising Genotypes of Black Gram [Vigna mungo (L.) Hepper]” was carried out at the Department of Biotechnology, Junagadh Agricultural University, Junagadh with objectives to analyse different 20 selected genotypes of black gram for molecular characterization using various PCR based molecular markers viz. Random Amplified Polymorphic DNA (RAPD), Inter Simple Sequence Repeats (ISSRs) and Simple Sequence Repeat (SSR). Along with, various nutritional and anti-nutritional parameters. The 20 selected genotypes of black gram were selected for the molecular study. Pooled study of molecular marker through RAPD, ISSR and SSR used to confirm the differences and similarity between the black gram genotypes. A total of 11 RAPD primers were amplified with total of 65 bands. There were 62 polymorphic bands out of 65 bands with average of 5.63 bands per primer. The percentage polymorphism obtained for RAPD primers is 94.37 %. The largest fragment of 3200 bp was amplified by CMN- A37 and the smallest fragment of 100 bp was amplified by OPL-03. The primer OPN-14 and CMN-18 amplified one- one unique band. 10 ISSR primers amplified a total of 43 bands. The ISSR marker UBC-811 produced maximum number of 7 bands. The primer UBC-807 amplified two unique band and UBC-825 amplified one unique band. The percentage polymorphism obtained for ISSR primers is 90 %. 12 SSR primers amplified 12 bands. Out 12 bands, 4 bands are monomorphic and 8 bands are polymorphic with 66.67 % polymorphism. All SSR markers produced single band. There was no unique band in SSR analysis. An average polymorphism information content (PIC) value for RAPD primer, ISSR primer and SSR primer were 0.87, 0.91 and 0.00, respectively. Primer index for RAPD, ISSR and SSR were 11.84, 14 and 1, respectively.
Open Access
RNA-SEQ FOR DIFFERENTIALLY EXPRESSED GENE(S) REGULATING DROUGHT AND SALT STRESS RESPONSE IN BARNYARD MILLET (Echinochola frumentaces L.) 3570
(JAU,JUNAGADH, 2022-09) VAGHELA TEJASVINI INDRAJITSINH; Dr. A. G. Vala; 2010120089
Barnyard millet (Echinochloa frumentaces L.) is grown for human consumptionas well as fodder.Barnyard millet is the second important small millet after finger millet having production and productivity 87 thousand tons and 857 kg/ha, respectively. Among small millets, it is the fastest growing millet and highly self-pollinated crop in to family Poaceae. Barnyard millets are high in fibre content, phosporous and calcium. The protein content in barnyard millet ranged from 11.1% to 13.9%. Barnyard millet grain contains about 65% carbohydrate, majority of which is in the form of non-starchy polysaccharide and dietary fibre. Barnyard has non-glutinous type of endosperm. The tannin content is at lowest level in barnyard millet (102.96 mg). Barnyard millet is highly suitable for commercial foodsfordiabetics, infants and pregnant women because of high iron content 38.5 to 42 ppm. Bardyard has low glycemic index and thus helps in type 2 diabetes, cardiovascular disease with regular intake of this millet.
Open Access
MOLECULAR CHARACTERIZATION, NUTRITIONAL AND ANTINUTRITIONAL PARAMETERS OF PROMISING MUNG BEAN [Vigna radiata (L.) Wilczek] GENOTYPES 3564
(JAU,JUNAGDH, 2022-09) TAVIYAD KRISHNABEN LAXMANBHAI; Dr. U. K. Kandoliya; 2010120085
The present investigation on “Molecular characterization, nutritional and antinutritional parameters of promising mung bean [Vigna radiata (L.) Wilczek] genotypes” was carried out at the Department of Biotechnology, Junagadh Agricultural University, Junagadh with objectives to analyse molecular characterization of 20 different mung bean genotypes using various PCR based molecular markers viz. Random Amplified Polymorphic DNA (RAPD), Inter Simple Sequence Repeats (ISSRs) and Simple Sequence Repeat (SSR) as well as to find out the phylogenic relationship among different mung bean genotypes. Also, estimation of nutritional and antinutritional parameters of mung bean genotypes was done. The 20 selected genotypes of mung bean were selected for the molecular study. Pooled study of molecular marker through RAPD, ISSR and SSR used to confirm the differences and similarity between the mung bean genotypes. Total 20 RAPD primers were screened, out of which 10 primers amplified a total of 63 bands. The per cent polymorphism obtained for RAPD primers was 75.09 per cent. The Polymorphism Information Content (PIC) values for RAPD markers were ranged from 0.93 to 0.98. For ISSR analysis, total 21 ISSR primers were screened, out of which 12 primers amplified a total of 75 bands. The per cent polymorphism obtained for ISSR primers was 79.26 per cent. The Polymorphism Information Content (PIC) values for ISSR markers were ranged from 0.80 to 0.98. For SSR analysis, total 15 SSR primers were screened, out of which 12 primers amplified a total of 18 bands. The per cent polymorphism obtained for SSR primers was 70.83 per cent. The Polymorphism Information Content (PIC) values for SSR markers were ranged from 0.00 to 0.88. Genetic similarity with all three molecular markers were determined for each pair of twenty mung bean entries which revealed that genotype GJM-2020 with GJM-2018 and GJM-2026 with Meha showed highest similarity (93%), like that GAM-5 and GJM 2017 showed lowest similarity (62%). Jaccard’s similarity coefficient and UPGMA method were used to develop a dendrogram which divided the genotypes into two main clusters A and B with an average resemblance of 68 %. The Main cluster A contained only one genotype (GJM-2017) which is most diverse among all the genotypes while Cluster B consist of all 19 Genotypes. The data generated from the present study may be
Open Access
DE NOVO TRANSCRIPTOME SEQUENCING AND METABOLOMIC PROFILING TO DISCOVER PUTATIVE GENES OF LITTLE MILLET (Panicum sumatrense L.) ASSOCIATED TO DROUGHT TOLERANCE 3624
(JAU JUNAGADH, 2022-10) DHAWALE RAMESH NARAYANRAO; Dr. R. S. Tomar; 1010119006
Little millet (Panicum sumatrense), having relatively diploid genome (2n=4x=36) with unknown genome size. It is one among the minor millets grown to a limited extent all over India up to altitudes of 2100 m. This crop is resistant to adverse agro-climatic conditions. The complex carbohydrates, phenolic compounds, antioxidants help to prevent metabolic disorders like diabetes, cancer, obesity etc., Although minor millets were superior to other cereals with many nutritional benefits due to its proximate composition of protein, fat, carbohydrates, ash, moisture, and energy, their utilization was limited because of low palatability, the coarseness of grain, and lack of diversified food preparations. Hence evaluation of the nutritional quality of millets would help to understand, diversifying the usage of millets in ensuring food, nutritional security in the ever-changing modern world. Keeping theses benefits in the mind present study was undertaken to find out putative genes and metabolites responsible for drought tolerance. The little millet genotype OLM 203/Tarini tolerant to drought was given in control and stressed condition in pots. 23 days old regularly watered seedling (control) and 32 days old water stressed seedlings were utilized for present experiment. In the current study, transcriptome analysis identified key genes regulating drought tolerance viz, in all 04-sample, number of reads varied 582,326 to 1,302,251 and average read length 497.13 to 589.32 bp. After trimming, there was decrease in number of reads, average read length and trimmed percentage which is varied between 569,157 to 1,265,909, 501.4 to 588.85, 90.74 to 97.74 % respectively. As well as, in all four samples number of total raw reads and average read length were 3,188,195 and 531.64 respectively and after trimming by CLC-GWB v20.0 number of raw reads decreased to 3,048,100 trimmed reads, trimmed read length 532.60, and trimmed percentage 95.24 %. Clean reads were obtained after quality control and they were subsequently used for De novo assembly and expression analysis. The generated draft assembly was 214498057 bp was approximately percentage of the total genome size of little millet is unknown. The maximum and minimum contig size was 40533 and 400bp in leaf and 398 base pairs in root respectively. However, the average size of contigs was 1138.04 bp and N50 was 1642 bp. Total de nono assembly were generated assembled reads of 270248626 to 214498057 with GC (%) was 48.39 from four samples with a hit length of 13112251 to 12759055 with GC (%) was 46.99. Differential expression analysis, total number of genes in comparison of 23 days leaf vs 32 days leaf (treated) were observed 7264, and in comparison of 23 days root vs 32 days root (treated) were extracted 7606. In expression profiling of 134 important DEGs, up or down regulated 20 important drought tolerance or resistance responsive genes in control and treated root of OLM-203 genotype was observed such as, in leaf XB34, and RS193 (up-regulated genes), WLIM1 (down regulated genes). In case of roots, SKI35 (up-regulated genes) while, MPK6 and TCMOp1 (down regulated genes). However, in leaf, up-regulated top 10 important drought tolerance or resistance responsive genes (LHW, CML25, PLP2, OBE1, SKI35p1, ATESY, SKI35p2, C3H7, PSAH1, PAP14) and in root MPK6, SPT41, THIC, LRX6, LRX5, COQ4, DMP5, LRX5, IAA6 and SERL2 genes were found to be switched on. Among these top 10 genes excluding SPT41, THIC (taking part in transcription process) genes takes part in drought tolerance in the genotype. There are top twenty commonly expressed drought tolerant or resistance genes in leaf and root of OLM-203 genotype viz., AX22A, BOB2, C3H32, C71B8, CAHC, CAL, CCS1, DGDG1, FLA2, GL52, GMPP2, HAK18, IAA6, IDL2, M3K1, NAC81, OHP1, PRX2C, SCP44 and TPR3. These all genes are responsive for positive regulator of stress signaling and drought tolerance mechanism. There are 36 up-regulated and 21 down-regulated serine transcripts were expressed in root and leaf of OLM-203 genotype of little millet respectively. ‘SKI35’ (Log₂ FC=7.66) up-regulated and ‘ALA2’ (Log₂ FC= -5.82) down-regulated genes are serine regulated differential expression genes in root of OLM-203 genotype. ‘NPRT1’ (Log₂ FC= 2.89) up-regulated and ‘KN7F’ (Log₂ FC= -4.11) down regulated genes are serine regulated differential expression genes in leaf of OLM-203 genotype. A perusal of dendrogram was clearly indicated that 134 drought tolerance genes were considered and three genes IAA6, ALA2, and HP301 among 134 genes played a key role during the drought or stress tolerance of the genotype. Most GO gene annotations of leaf samples were categorized as having ‘biological process’ (35%), followed by ‘cellular component (19%) and ‘molecular function’ (46%). Within the top represented category of ‘cellular process (125 contigs)’ were enriched, as was ‘metabolic process’ (117 contigs) and the next most enriched ortholog-biosynthesis of ‘cellular component’ (150) found in this category. Most GO gene annotations of root samples were categorized as having ‘biological process’ (573 contigs), followed by ‘molecular function (401 contigs) and ‘cellular components (166 contigs)’. In root samples, 1140 genes were found which are most enriched with biological process, molecular function and cellular component, whereas in leaf samples 670 genes most enriched ortholog-biological process, molecular function and cellular component. Enzyme commission (EC) numbers were assigned for control leaf vs treated leaf with 105 enzyme codes for 692 unique sequences, whereas in case of control root vs treated root with 105 enzyme codes for 699 unique sequences. Among all the leaf samples, the GO level distribution graph showed 1270 total annotations with mean level 6.036%, and having standard deviation 2.509, whereas in root samples containing 1617 total annotations with mean level 5.947%, and having standard deviation 2.516. Several putative functions were identified functional domains such as zinc finger, drought tolerance, serine related proteins. The confirmation results of repeats obtained by STRING 10.0 shown in MCL clustering of repeats depicted the 11 gene clustered with the specific repeats obtained from STRING 10.0. Six protein sequences (genes) viz., IPR019734 Tetratricopeptide, IPR013105 Tetratricopeptide, IPR018108 Mitochondrial subs, IPR019734 Tetratricopeptide string MCL clusters, IPR013105 Tetratricopeptide string MCL clusters, IPR002885 Pentatricopeptide string MCL clusters deciphered the role of its genes were regulated by their 11 genes cluster jointly. While, Venn diagram showing a total of 134 genes were found commonly shared and unique DEGs in leaf and root and 27 up-regulated genes were found commonly in leaf and root. There are 18 enzymes were analyzed and showed 44 KEGG pathways were found to be regulated. There were unique and common sequences regulate the 53 pathways. There are 35 unique and common KEGG pathways regulated by 58 enzymes of OLM-203 genotype. Total 147 KEGG pathways were commonly operated among control and treated samples of root and leaf. There are 15 and 35 number of unique and common KEGG pathways regulated by specific enzyme and sequences in root and leaf of genotype respectively up to 32 days of the genotype. Using the NIST library for the metabolomics study, metabolites were identified as sugars, sugar alcohols, sugar acids, fatty acids, and others such as dicarboxylic acid, diterpene alcohol, organic acid, and sugar amine. The detailed of total number of sixty (60) metabolites produced in both samples. In the mean decrease accuracy, metabolites were linolenic acid, mannonic acid, hexadecanoic acid, pentonic acid, glucopyranosid, glucohexodialdose, and malic acid were present in higher concentrations in the OLM-203 genotype. In treated samples, glucose, threonine, acetamide, serine, oleic acid, fructose glucose oxime, and hydroquinone were intensively observed, while in control sample, linolenic acid, mannonic acid, hexadecanoic acid, pentonic acid, glucopyranosid, glucohexodialdose, and malic acid were observed in OLM-203. Sugar also acts as a signaling molecule and helps to modulate the plant’s growth, development, and response to multiple stresses. Mannose was found to be accumulated in the leaf of OLM-203 (Tarini) in stress conditions; similar findings were reported earlier in the drought-tolerant little millet. Genomic and metabolomics changes were observed during drought treatment in tolerant genotype ‘OLM-203/Tarini’. It was observed that the typical increase of stress-related genes and metabolites. Therefore, it can sum-up that ‘OLM-203/Tarini’ genotype was well tolerated genotype and functions was normal even under water stress, although the exact complex mechanism of tolerance remains to be investigated. There were 128 EST-SSR identified which serves as a resource of high quality transcripts for gene discovery and development of functional markers associated with water stress tolerant. In the present study, sequence collection represents a major transcriptomics resource for little millet genotype and the large number of genetic markers predicted should contribute to future research thrust.
Open Access
TRANSCRIPTOME AND HORMONE PROFILING OF GROUNDNUT (Arachis hypogaea L.) TO UNDERSTAND THE MECHANISM OF FRESH SEED DORMANCY 3441
(JAU,JUNAGADH, 2021-12) HEMANGINI AMBELAL CHAUDHARI; Dr. M. K. Mahatma
Cultivated groundnut (Arachis hypogaea), is widely grown throughout the world, used for food, fodder and for industrial purpose and can be easily grown by resource poor farmers. Its genome sequencing is completed hence related information like gene discovery; marker development and even gene expression study have been carried out at vivid platform. India is one of the largest producers of groundnut in the world. Precocious germination of the seed of groundnut crop in to the field due to rainfall during harvesting time can causes huge yield loss and can affect the quality of seeds. Therefore, present study was carried out to understand molecular mechanism of fresh seed dormancy in groundnut. To fulfill our objectives transcriptome analysis along with phytohormone and protein profiling was carried out in highly dormant (d; NRCG 14380) and non-dormant (nd; TAG 24) genotype at different seed germination stages
Open Access
COMPARATIVE ANALYSIS OF POTATO FARMING: CONTRACT vis-a-vis CONVENTIONAL 3519
(JAU, 2022-08) PATEL PRIT DINESHBHAI; PATEL PRIT DINESHBHAI
Open Access
MICROBIOME DIVERSITY AND METAGENOMICS ANALYSIS ASSOCIATED WITH FUSARIUM WILT IN CUMIN RHIZOSPHERE 3512
(JAU, 2022-08) HIRANI NIMISHABEN VINUBHAI; Hirani Nimishaben Vinubhai Dr. H. P. Gajera; HIRANI NIMISHABEN VINUBHAI
Cumin (Cuminum cyminum Linnaeus) belongs to family Apiaceae and an
Open Access
TRANSCRIPTOME SEQUENCING AND METABOLOMIC CHARACTERIZATION OF BARNYARD MILLET (Echinochloa frumentacea L.) TO DISCOVER PUTATIVE GENES INVOLVED IN SPIKE DEVELOPMENT AND ITS NEUTRACEUTICAL PROPERTIES
(JAU,JUANAGDH, 2020-12) PADHIYAR SHITALBEN MALDEBHAI; Dr. R. S. Tomar
Barnyard millet comprises two different cultivated species, Japanese barnyard millet (Echinochloa esculenta L.) and Indian Barnyard millet (Echinochloa frumentacea L.) belongs to the plant family Poaceae.The other names of barnyard millet in Gujarati is moraiyo. Barnyard or sawa millet is the fastest growing crop of all millet, which can produce ripe grains within 45 days from the sowing time under optimal weather conditions. In addition to these agronomic advantages, the grains are valued for their high nutritional value and lower expense as compared to major cereals like rice, wheat, and maize. It is important minor millet in Japan, China, India and other South East Asian countries. Barnyard millet is a multi-purpose crop which is cultivated for food and fodder. Barnyard millet grains are a rich source of dietary fiber, iron, zinc, calcium, protein, carbohydrate, magnesium, fat, vitamins, some essential amino acids and, most notably, contains more micronutrients (iron and zinc) than other major cereals. Due to low glycemic index and high dietary fiber, it helps in preventing diabetes and cardio vascular disease with regular intake. Barnyard millet could be a good source of iron for vegetarians. Keeping these benefits in the mind present study was undertaken to find out candidate genes and metabolites responsible for Fe content in barnyard millet genotypes. In the current study, biochemical parameter (proximate and minerals) was collected out in 30 barnyard millet genotypes and result shown that total carbohydrate content was found in range between 53.51-73.98 %. The range for total protein (9.55- 11.99 %), crude fiber (11.14–16.64 %), moisture content (4.47-7.68 %), ash content (3.47 – 5.99 %), total fat content (3.38 - 6.95% ), energy (304.45–380.11 Kcal/mole), fatty acid profiling reveled Oleic acid content was observed higher (81.19 %) followed by Palmitic acid (69.33 %), Linolenic acid (50.96 %) and Linoleic acid (48.11 %), nitrogen (1.56-2.72 %), phosphorus (326.67-486 mg/100g), potassium (170-383 mg/100g), sodium (31.67-51 mg/100g), iron content (3.83-11.14 mg/100g), zinc (2.01- 3.88 mg/100g), manganese (1.33-2.85 mg/100g), calcium (35.83–209.18 mg/100g), copper (0.161-0.407 mg / 100g), magnesium (101.65 and 182.21 mg/100g) and boron (0.45-1.99 mg /100g) were observed in 30 barnyard millet genotypes. Two barnyard millet genotypes having high Fe (BAR-1433) and low Fe (BAR 1423) in their seeds were taken for transcriptome and metabolomic analysis during spike development stage. Total pool 30 HQ reads were considered for de novo assembly optimization using four standard assembler Trinity, Soapdenovo_trans, CLC and CAP3 assembler. The trimmed number of reads yielded a total of 138 million high-quality reads, which were assembled by trinity into 4,88,689 transcript. The trinity generated highest transcript with assembly size of 340 MB, N50 size of 1202 and N90 size of 611. CAP3 assembler was employed to reduce transcript redundancy and it generate 27,228 transcript with assembly size of 31.84 MB, N50 size of 1570 bp and N90 size of 723 bp. Moreover, more redundancy were removed by CD-HIT programme and assembler which generated 20,849 transcripts with size assembly of 24.42 MB, N50 size of 1604 bp and N90 size of 719 bp. Transcriptome analysis identified key genes regulating Fe accumulation like CIAO1, ISAM1 and UCRIA during different spike development stage. Further, Ferritin, ABC, Cytochrome, Metal, Zinc, Serine, Wall-associated and Cytochrome, Mitogen and Photosynthesis regulating differential gene also were identified which are also involved in iron content of Low Fe Vs High Fe barnyard millet genotype. Functional annotation was carried out for Gene Ontology (GO) enrichment and KEGG pathway enrichment from significant DEGs. Gene ontology (GO) and pathways analysis revealed that different metabolic pathway like glycolysis, purine and pyruvate metabolism based on expressed genes revealed the mechanism of minerals transportation. The resources generated in this study will facilitate discovery of new genes and further EST-SSR markers and thereby accelerate both genetic and functional genomic research in barnyard millet. Identified genes may be used for marker-assisted selection and breeding to develop minerals rich crops. The Comparison of transcript expression levels between transcriptome data and qRT-PCR depicted positive correlation. The validation through qRT-PCR was carried out using ACBL, ABA, RP8L, PDF1 and PHY which were among the up and down regulated transcripts in different stages of barnyard millet. Metabolome profiling of barnyard millet genotypes containing high Fe and low Fe was performed using GC-MS platform and results observed highest percentage of sugar and sugar alcohol (34%) followed by organic acid (26%), Amino acids, sterol, other compound (23%) and fatty acid (17%) in spike development stages of Low Fe Vs High Fe genotypes. Heat map revealed that during spike emergence the metabolic compounds like d-Ribose, D-Fructose, D-Glucose, Galactose, D-Turanose, Glucopyranose, D-Mannitol, Hexadecanoic acid, Docosanoic acid, alpha.- Glycerophosphoric acid and .beta.-Sitosterol were found high in high Fe genotype in comparison to Low Fe genotype. The hierarchical cluster analysis, revealed that high Fe five spike development stages shares close metabolite pool to each other and in low Fe four spike development stages also shares similarly metabolite pool to each other except spike emergence stage of Low Fe genotype. PCA showed that the expression patterns of the five developmental stages differed significantly.

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