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Anand Agricultural University, Anand
Anand Agricultural University (AAU) was established in 2004 at Anand with the support of the Government of Gujarat, Act No.(Guj 5 of 2004) dated April 29, 2004. Caved out of the erstwhile Gujarat Agricultural University (GAU), the dream institution of Sardar Vallabhbhai Patel and Dr. K. M. Munshi, the AAU was set up to provide support to the farming community in three facets namely education, research and extension activities in Agriculture, Horticulture Engineering, product Processing and Home Science. At present there seven Colleges, seventeen Research Centers and six Extension Education Institute working in nine districts of Gujarat namely Ahmedabad, Anand, Dahod, Kheda, Panchmahal, Vadodara, Mahisagar, Botad and Chhotaudepur
AAU's activities have expanded to span newer commodity sectors such as soil health card, bio-diesel, medicinal plants apart from the mandatory ones like rice, maize, tobacco, vegetable crops, fruit crops, forage crops, animal breeding, nutrition and dairy products etc. the core of AAU's operating philosophy however, continues to create the partnership between the rural people and committed academic as the basic for sustainable rural development. In pursuing its various programmes AAU's overall mission is to promote sustainable growth and economic independence in rural society. AAU aims to do this through education, research and extension education. Thus, AAU works towards the empowerment of the farmers.
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ThesisItem Open Access LABORATORY STUDIES ON BIOLOGY AND FEEDING POTENTIAL OF MEXICAN BEETLE, Zygogramma bicolorata Pallister ON PARTHENIUM, Parthenium hysterophorous L. AND EVALUATION OF HERBICIDES FOR THEIR SAFETY TO THE BIOAGENT(AAU, Anand, 2013) PAWAR, SATISH RAMCHANDRA; Korat, D. M.Investigations on biology, morphometries and feeding potential of Mexican beetle, Zygogramma bicolorata Pallister (Chrysomelidae: Coleoptera) and toxicity of weedicides as well as effect of temperature on biological attributes of the bioagent were carried out in Biological Control Research Laboratory, Anand Agricultural University, Anand (Gujarat) during theyear 2011 to 2012. Studies on biology of Z. bicolorata on Parthenium hysterophoms L. revea:ied that the female laid their eggs either singly or in cluster on under surface of leaves. Eggs were oblong in shape, slightly elongated, smooth and the surface was finely reticulated. There were four distinct larval instars. Newly hatched larvae were yellowish in colour and gradually turned in creamy white with the advancement of age. Adults of Z. bicolorata were elongate and oblong in shape. Dorsal surface was strongly convex and glabrous. In general, females found relatively larger in size than males. Average egg, larval, pupal and adult period was 3.93+0.80, 13.87 ± 1.36, 7.40 ± 1 . 1 8 and 32.40 ± 8.05 (males) to 44.53 + 7.33 (females) days, respectively. Entire life-span of female and male completed in 59.13 ± 7.75 and 71.33 ± 8.78 days, respectively. Fecundity, hatching percentage, adult emergence percentage and male to female sex ratio was 669.73 ± 141.34 eggs, 55.75 ± 12.27%, 76.67 ± 14.82% and 1 : 1.26, respectively.ThesisItem Open Access EXPRESSION PROFILING, SNP DETECTION AND VALIDATION IN SQUAMOUS CELL CARCINOMA OF HORN IN KANKREJ CATTLE (Bos indicus) USING NEXT GENERATION SEQUENCING(AAU, Anand, 2014) KORINGA, PRAKASHKUMAR G.; Joshi, Chaitanya G.Horn cancer is a widely prevalent cancer amongst Kankrej cattle (Bos indicus) seen sporadically, especially in case of working class of castrated male animals i.e. bullocks. A transcriptome envisaged characterization as well as correlation to known genomic changes such as structural and copy number alterations, focused ins/dels and single nucleotide mutations. Here, we employed high throughput RNA-seq using GS-FLX Titanium for characterization and comparison of normal and cancerous horn transcriptome in Bos indicus. A total of 909,362 reads with average read length of 405bp for horn cancer (HC) and 583,491 reads with average read length of 411bp for horn normal (HN) were obtained by sequencing gene transcripts derived from HC and HN tissues. Assembled data were analyzed for identifying novel as well as differentially expressed transcripts using CLC Genome Workbench. RNA-seq analysis using different bioinformatics pipelines and software identified differentially expressed genes i.e. upregulation of KRT6A, KRT6B, KRT6C, KRT14, SFN, KRT84, PI3, CAl, C0L17A1, ANLN, SERPINB5 etc., as well as down-regulation of NR4A1, FOSB, LRIGl, BOLA, SCGBIAI, CXCL17, KRT19, BPIFBl, NR4A1 and TFF3 etc., in HC tissues. The signaling pathway investigation in this study revealed many of the cancer related pathways which mainly include cell cycle regulation pathways, p53 tumor suppressor pathways, NFKB and MAPKs pathways, LPS signaling pathway and PI3K-Akt pathways. The resuh of transcriptome expression profiling was validated using RT-qPCR in nine randomly selected genes. It revealed concordance of gene expression profile with RNA-seq analysis. We also used transcriptome data to elucidate complexity of the alternative splicing in HC transcriptome. We identified potential candidate splice variants that might be helpful in development of relevant biomarkers for early diagnosis of HC. The fiiture studies targeted at in depth characterization of these potential candidate splice variants might change the currently used clinical approaches. Herein we characterized global landscape of alternative splicing events exhibited by pair of HC and HN tissue and confirmed selected alternative splicing events with significant association to HC by RT-qPCR. Ine analysis of the same RNA-seq data using SeqMan Pro Version 10.0.0 resulted in to a 9532 and 7065 SNPs as well as 1171 and 1172 Indels in HC and HN, respectively. Out of total, 7889 SNPs and 1736 Indels uniquely present in HC, 5886 SNPs and 1146 Indels uniquely present in HN are novel and reported first time in Bos indicus, whereas rest are already reported in Bos taurus dbSNP database at NCBI. The gene-associated SNPs and Indels were high in upregulated genes of HC as compared to HN tissues. SNPs identified in RNA-seq analysis were validated in fiirther studies in two groups consisting of 50 animals each of HC and HN bullocks. DNA from HC tissue and blood of HN individual was extracted and 96 pairs of primers were used to generate amplicons of an average 300bp to get sequenced using Ion Torrent PGM. The resulting reads were assembled using SeqMan N Gen of DNASTAR and data were analyzed using Arraystarll. Case control analysis was carried out to find SNP significantly associated with HC. SNP at position 63251805 (dBSNP ID rsl36870681) identified in BPIFAl can serve as a potential candidate genetic marker in HC. The SNPs and Indels identified in this study will be useful resource for future studies to understand genetic basis for phenotypic variation between Bos taurus and Bos indicus as well as cancers in animals. A very large number of SNPs are essential for the designing and construction of arrays. SNPs identified in this study will enrich the dbSNP database of NCBI (http://www.ncbi.nlm. nih.gov/projects/SNP/) and will be useful resource for array designing. This study is the first attempt to reveal novel transcripts, differentially expressed genes as well as identification and validation of SNPs using digital expression analysis in Bos indicus and provides novel insights into bovine transcriptome. Our study will serve as a step further in detailed characterization of HC transcriptome and provide firm base to explore and mitigate HC at finer resolution. The present findings would provide basis for further screening of genes and identification of markers for early diagnosis and therapeutic intervention of HC.ThesisItem Open Access HAEMATOLOGICAL, BIOCHEMICAL AND ENDOCRINE PARAMETERS AT DIFFERENT AGES AND PHYSIOLOGICAL STAGES IN GIR CATTLE AND JAFFARABADI BUFFALOES(AAU, Anand, 2012) JACOB, NINAN; ARYA, J. S.The study titled "Haematological, Biochemical and Endocrine parameters at different Ages and Physiological stages in Gir cattle and Jaffarabadi buffaloes" was imdertaken in different age groups and physiological stages in Gir cattle and Jaffarabadi buffaloes with the objective to determine and compare the species differences for (i) haematological parameters viz. TEC, Hb, PCV, TLC, DLC (ii) biochemical parameters viz. plasma glucose, total protein, albumin, total cholesterol, HDL cholesterol, triglycerides, urea, uric acid, creatinine, calcium, phosphorus, magnesium, sodium and potassium (iii) enzymes viz. aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, acid phosphatase, and lactate dehydrogenase (iv) hormonal levels of triiodothyronine, thyroxine, insulin, growth hormone, Cortisol, estradiol - 17 β, progesterone and testosterone and (v) to observe the milk components viz. percentage of fat, protein and lactose in lactating Gir cows and Jaffarabadi buffaloes and their relationship with the stage of lactation under study. The blood samples were collected from Gir and Jaffarabadi females (n=8 for each sampling stage) at 1 wk, 1, 3, 6, 12, 24 and 36 months age, at 1, 2 and 3 month of lactation and in non-lactating pregnant and non-pregnant animals. In Gir and Jaffarabadi males (n=6 for each sampling' stage) blood samples were collected at 1 wk, 1, 3, 6 and 12 month of age and in bulls. Castrated males were also used for blood sampling in Gir males. A total of 270 blood samples were collected and the analysis was conducted by standard techniques.ThesisItem Open Access MONITORING POSTPARTUM REPRODUCTIVE PERFORMANCE IN KANKREJ COWS THROUGH CLINICAL DIAGNOSIS, BLOOD PROFILE AND HORMONAL THERAPY(AAU, Anand, 2012) NAIKOO, MEHRAJUDDIN; Dhami, A. J.The present study was carried out at Livestock Research Station of the University on 42 Kankrej cows of 2nd to 4th parity. The chief objectives were: to monitor the early postpartum period (0-90 days) clinically and through plasma profile of progesterone, metabolites and macro-micro minerals at 10 days intervals; to evaluate the efficacy of a sustained release mineral supplement (Mega bolus PO) on the day of calving and five oestrus induction and synchronization protocols (Ovsynch, CIDR, Ovsynch + CIDR, Cosynch and PGF2α) on day 90-95 postpartum towards augmenting reproductive efficiency of anestrous and subestrous cows (6 animals in each group), keeping 6 normal cyclic animals as control, and its effect on above profile till day 40 post-treatment/post- AI, and to compare plasma profiles of conceived and non-conceived cows at first Al. The time required for expulsion of fetal membranes, weight of expelled fetal membranes and the birth weight of calf (pure and crossbred) were 5.04 ± 2.0 hrs, 2.84 ± 0.76 kg and 24.29 ± 1.54 kg, respectively. The Kankrej cows showed complete uterine involution by mean interval of 36.80 ±1.21 (range 32-45) days postpartum. The interval for occurrence of first oestrus postpartum clinically and through plasma P4 profile was 105.49 ± 1.66 (range 86-106) and 56.42 ± 3.88 (range 30-80) days, respectively (P<0.05). The first service and overall conception rates obtained at spontaneous/ induced oestrus, within 150 days postpartum were 30.95 (13/42) and 40.47 (17/42) per cent. The comparative evaluation of the efficacy of five oestrus induction/ synchronization protocols tested, on 6 cows each, viz. Ovsynch, CIDR, Ovsynch + CIDR, Cosynch and PGF2α revealed oestrus induction response of 66.66, 83.33, 50.00, 66.66 and 66.66 per cent, respectively, with behavioural signs at FTAI as confirmed by palpation per rectum. The first service conception rates obtained were 16.66, 33.33, 16.66, 50.00 and 50.00 per cent, respectively, as compared to 33.33 per cent in normal cyclic control cows. The corresponding second service conception rates were nil, 25.00, 20.00, nil, nil and nil per cent, as compared to 25.00 per cent in untreated control animals. The overall conception rates of three cycles over the 45 days period were 33.33, 50.00, 33.33, 50.00 and 50.00 per cent, respectively, as against 50.00 per cent in normal cyclic group. The results of CIDR, Cosynch and PGF2α protocols were better than the Ovsynch and normal control groups.ThesisItem Open Access EFFECT OF PROGESTERONE AND HEAT STRESS ON REPRODUCTIVE PERFORMANCE OF DAIRY COWS AND VALIDATION OF CONTINUOUS BODY TEMPERATURE MEASUREMENT IN IDENTIFYING ESTRUS(AAU, Anand, 2012) SUTHAR, VISHAL S.; Dhami, A. J.The present study with four experiments and manifold objectives were conducted on German (HF) dairy cows at or affiliated farm facilities of Clinic for Animal Reproduction, Freie Universitaet Berlin, Germany, during January 2010 to June 2012. The overall objectives of this study were to evaluate: 1) performance of temperature data logger in in vitro and in vivo conditions, 2) validity of BT to identify induced estrus, 3) effect of exogenous and endogenous P4 on BT of dairy cows and 4) and to study effect of heat stress on BT and postpartum performance of dairy cows. In first study to evaluate performance of temperature data logger (Minilog 8, Vemco Ltd., Halifax, Canada), three sub-experiments were conducted. The study began with an in vitro validation of 24 temperature loggers comparing them to a calibrated liquid-in-glass thermometer as a reference method (sub-experiment 1). The association and agreement between the 24 temperature loggers with the reference method was r = 0.996 (P < 0.001) and a negligible coefficient of variance (0.005) between the loggers. In vivo temperature loggers were tested in 11 healthy postpartum cows (sub-experiment 2) and 12 early postpartum cows with greater BT (sub-experiment 3). Temperature loggers were set to record VT and RT at 1 min intervals. To prevent rectal and vaginal straining and potential expulsion of temperature logger an epidural injection of 2.5 ml of 2% Procain was administered. Association between RT and VT was r = 0.92 (P < 0.001) in sub-experiment 2 and r = 0.94 (P < 0.001) in sub-experiment 3 with a negligible difference of -0.1 and 0.01°C, respectively. Bland-Altman plots demonstrated an agreement between RT and VT for healthy and early postpartum cows with greater BT in sub-experiment 2 and 3, respectively. Therefore, continuous VT monitoring with temperature loggers can be used as a measure for BT in dairy cows.ThesisItem Open Access REGULATION OF ACTIVIN TYPE II RECEPTOR B (ACVR2B) EXPRESSION THROUGH RNA INTERFERENCE IN GOAT MYOBLAST CELLS(AAU, Anand, 2014) PATEL, AMRUTLAL KALUBHAI; Joshi, Chaitanya G.Enhancement of skeletal muscle mass through genetic manipulation has drawn attention to increase the meat production in the farm animals. Among the various techniques of regulating gene expression, RNA interference (RNAi) has been proposed as a promising tool to suppress the target gene expression. Attempts have been made to increase the skeletal muscle mass in transgenic animals through knockdown of Myostatin, a gene with potential negative effect on muscle growth. It has been well established that myostatin mediates its action through binding to its cell surface receptor mainly to activin type II receptor B (ACVR2B). Besides regulating myostatin activity, ACVR2B has also been known to regulate the activity of other Transfonning Growth Factor beta (TGF-(3) superfamily ligands which negatively regulates muscle growth. The inhibition of ACVR2B signaling has shown dramatic increase in the muscle mass to a greater extent than myostatin inhibition. Hence, in the present study we aimed to investigate the possibility of ACVR2B knockdown to enhance the myogenesis in goat through various RNAi methods such as expression of short hairpin RNAs (shRNAs) under U6 and CMV promoters and expression of artificial microRNAs (amiRNAs) under CMV and muscle specific muscle creatine kinase (MCK) promoters. Further we studied effect of ACVR2B knockdown on the expression of myogenic regulators and assessed induction of undesired interferon response against RNAi vectors. Among the seven shRNAs tested, the U6 promoter driven shRNAs showed 63 (p= 0.0004), 76 (p= 0.0001), 75 (p=0.0000), 74 (p= 0.0005), 80 (p= 0.0001), 74 (p= 0.0000) and 57% (p= 0.0013) silencing with sh1 to 7, respectively in HEK293T cells whereas 24 (p= 0.1497), 24 (p= 0.2243), 15 (p= 0.3988), 31 (p= 0.1263), 14 (p= 0.4425), 46 (p= 0.0318), and 26 % (p= 0.1288) silencing of endogenous ACVR2B with shl to sh7, respectively and 53 (p- 0.0005), 32 (p= 0.0171), 38 (p= 0.0025), 66 (p= 0.0002) and 51% (p= 0.0008) with sh3, sh4, sh5, sh6 and sh7, respectively against ectopically expressed goat ACVR2B in goat myoblasts ceils. The knockdown of endogenous ACVR2B resulted in the 116, 105, 84, 64, 119, 102, 121. and 157% expression of MyoD; 131, 128, 128, 123, 104, 103, 69, and 157% of MyoG in sh1 to sh7 transfected cells, respectively. The transfection of U6 driven shRNA resulted in the induction of OAS1, a marker for innate interferon response by 3 to 1861 fold in 293T cells and up to 94 fold in the goat myoblasts cells. In an attempt to overcome the undesired cellular toxicity associated with U6 driven shRNAs as reported in the number of studies, we expressed these shRNAs under CMV promoter. The CMV driven shRNAs showed weak silencing of 37 (p= 0.1622) and 18% (p= 0.4877) by sh1 and sh3, respectively in HEK293T cells whereas 7% (p= 0.5749) by shl and 4% (p= 0.7493) by sh5 in goat myoblasts cells. Unlike suggested earlier, we observed significant induction of interferon response to CMV driven shRNAs up to 46 fold in 293T cells and 105.3 fold in goat myoblasts cells. Alternatively, we assessed another RNAi approach using amiRNAs which mimics the endogenous miRNA biogenesis pathway. Among the four amiRNAs tested by placing them in 5'-UTR region of GFP reporter, we observed 64 (p=0.0004), 77 (p=0.0002), 1 (p=0.8712), and 41% (p=0.0115) silencing in 293T cells by ami204, ami318, ami735 and ami878, respectively against exogenously expressed goat ACVR2B; 19 (p=0.3593) and 9% (p=0.4977) by ami204 and ami318, respectively against endogenous ACVR2B and 23% (p=0.0444) by ami318 against exogenously expressed ACVR2B in goat myoblasts cells. Since, amiRNAs placed in 5'-UTR were shown to affect the translation of reporter GFP, we further placed them in 3'-UTR of GFP which resulted in enhanced expression of GFP thereby enabling the monitoring of expression of amiRNAs. The 3'-UTR derived amiRNAs showed 50% (p=0.0002) silencing only by ami3I8 in 293T cells whereas 47 (p=0.0193), 16 (p=0.2959), 19 (p=0.1547), and 28% (p=0.0770) by ami204, ami318, ami735 and ami878, respectively against endogenous and 67%) (p=0.0004) by ami318 against overexpressed ACVR2B in goat myoblasts cells. The expression of myogenic regulators MyoD remained unchanged by amiRNAs cloned in the 5'-UTR whereas expression of MyoG was significantly up regulated by ami878 (p=0.0089). However, amiRNAs cloned in 3'-UTR showed significant down regulation of MyoD by 51 (p=0.0007), 27 (p=0.0232), 29 (p=0.0074), and 31% (p=0.0104) and MyoG by 36 (p=0.0034), 12 (p=0.2532), 22 (p=0.0303), and 37% (p=0.0026) by ami204, ami318, ami735 and ami878, respectively. As observed for U6 and CMV driven shRNAs, CMV driven amiRNAs showed significant induction of interferon response in 293T (up to 121.7 fold) and myoblasts (212.5 fold) cells. As ACVR2B has been shown to be essential for embryonic development, we tested the possibility of its knockdown in skeletal muscle using muscle specific MCK promoter. Among the MCK and MSTN promoters with and without two repeats of MCK enhancer, we observed maximum transcriptional activity by MCK promoter in goat myoblasts cells. We thus tested best amiRNAs (ami204 and ami318) by expressing under MCK promoter which showed 22% silencing efficacy by ami318 in 293T cells and 32% silencing efficacy in goat myoblasts cells by transient transfection assay. Further to test the possibility of ACVR2B knockdown after stable integration of amiRNAs into goat myoblasts genome, we generated lentivirus particles carrying amiRNAs expression cassettes and transduced the goat myoblasts. The myoblasts cells stably integrated with amiRNAs showed ~8% silencing by ami318 which was increased to 34% upon induction of differentiation under muscle specific promoter. Western blot analysis revealed 41% and 57% silencing by 5'-ami204 and 5'-ami318, respectively whereas 14% and 35% silencing by 3'-ami204 and 3'-ami318, respectively in stable myoblasts upon induction of differentiation. Unlike transient transfection assay vv'hich showed positive correlation of expression of ACVR2B with myogenic regulators, its stable knockdown resulted in up regulation of MyoD in 5'-UTR derived ami204 and ami318 and overall down regulation of MRFs in 3'-UTR derived ami204 and ami318 integrated goat myoblasts cells. The 5'- UTR derived ami204 and ami318 showed increased rate of cell proliferation as well as myoblasts fusion in stable goat myoblasts compared to scramble control indicating growth promoting effect of ACVR2B knockdown. The skeletal muscle specific partial knockdown of ACVR2B is unlikely to affect the embryonic survival and it will be interesting to further assess its possible growth promoting effect in adult animal by generating transgenic goat through somatic cell nuclear transfer of goat myoblast cells stably integrated with amiRNAs.ThesisItem Open Access EFFECT OF ROW RATIOS, PHOSPHORUS LEVELS AND WEED MANAGEMENT PRACTICES ON PERFORMANCE OF MUSTARD - CHICKPEA INTERCROPPING AND ITS RESIDUAL EFFECT ON SUMMER BLACK GRAM UNDER MIDDLE GUJARAT CONDITIONS(AAU, Anand, 2013) GAIKWAD, VAIBHAV POPAT; Patel, J. J.An experiment was conducted during rabi season in the year 2010-2011 and 2011-2012 with succeeding summer black gram at College Agronomy Farm, B. A. College of Agriculture, Anand Agricultural University, Anand, Gujarat to study the "Effect of row ratios, phosphorus levels and weed management practices on performance, of mustard - chickpea intercropping and its residual effect on summer black gram under middle Gujarat conditions" The soil of the experimental plot was loamy sand in texture. It was low in organic carbon and available nitrogen, while medium in available phosphorus and high in available potash with pH 7.9 and EC value 0.11dsm-1. There were thirty treatment combinations comprised of five row ratios IC1 Chick pea (Sole), IC2 Mustard (Sole), IC3 Mustard + Chick pea (1: 1), IC4 Mustard + Chick pea (1: 2), IC5 Mustard + Chick pea (1: 3) and two levels of P 1 (25 kg P2O5 ha-1 ) , P2 (50 kg P2O5 ha-1) and three levels of weed management practices ( W1 :Weedy check , W2 : Hand weeding at 20 and 40 DAS, W3 : Pendimethalin @ 1 kg ha-1 (Pre-emergence) tested in randomized block design under factorial concept with three replications. Plant population of mustard recorded at 15 DAS and at harvest showed non-significant differences due to different row ratios. The plant heights at all the crop growth stages were significantly influenced due to different row ratios. Higher plant height (30.89 cm) was recorded under the treatment IC5 Mustard + Chick pea (1:3) row ratio while significantly the lowest plant height (27.94 cm) at 30 DAS was observed under the treatment IC3 Mustard + Chick pea (1:1). The difference in the plant height recorded at 60 DAS due to the different row ratios was non-significant on pooled basis. While plant height recorded at 90 DAS and harvesting was significantly influenced due to different row ratios. Significantly the highest plant height (204.83 and 213.94 cm) were recorded under the treatment IC2 (sole mustard) at 90 DAS and at harvest during second year and on pooled basis, respectively. Lower plant height (193.31 and 207.44 cm) of mustard were recorded under the treatment IC5 Mustard + Chick pea (1:3) which was remained at par with the treatment IC3 Mustard + Chick pea (1:1) and IC4 Mustard + Chick pea (1: 2) at harvest, respectively on pooled basis. The result revealed that different row ratios did not exert its significant influence on number of siliquae per plant and length of siliqua on pooled basis analysis. The results pertaining to seed and straw yield, harvest index and test weight revealed that the different row ratios registered significantly higher mustard seed and straw yield, harvest index as well as test weight were observed under the treatment IC3 Mustard + Chick pea (1: 1). Lower mustard seed yield, straw yield, harvest index as well as test weight were observed under the treatment IC5 Mustard + Chick pea (1:3). Plant height of chickpea recorded at 25, 50 DAS and at harvest was influenced significantly by the different row ratios, respectively. It had non-significant effect on pooled basis result at 25 DAS and at harvest. Plant heights measured at 50 DAS as influenced by the different row ratios were significant during both the years and on pooled basis. Significantly the maximum plant height (41.81 cm) of chickpea was observed under the treatment IC3 Mustard + Chick pea (1: 1) on pooled result. It was remained at par with the treatment IC4 Mustard + Chick pea (1:2). Significantly the lowest plant height (33.72cm) was recorded under treatment IC] sole chick pea. Among the various row ratios, the treatment IC3 Mustard + chickpea (1:1) recorded significantly the highest number of pods per plant (53.55), number of seeds per pods (1.67) seed (809 kg ha-1), dry gotar yield (1678 kg ha-1), harvest index (41.1 %) and test weight (25.63 g.) of chickpea on pooled basis, respectively. Significantly less number of pods per plant (27.76), seeds per pod (1.52), seed (300 kg/ha) and dry gotar yield (502 kg ha-1) of chickpea were recorded under the treatment IC5 Mustard + Chick pea (1: 3) on pooled basis. While significantly the lowest harvest index (25.9 %) and test weight (24.7 g.) was observed under the treatment IC1 (sole chick pea), respectively. As far as interaction effect concern all the possible interaction of different factors had the significant influence on the harvest index of mustard. Different phosphorus levels did not differ significantly for plant height on pooled basis at 30, 60, 90 DAS and at harvest while also it exerted non-significant effect on the yield attributes of mustard (number of siliquae per plant, length of siliqua, test weight of mustard and oil content) on pooled basis.ThesisItem Open Access COMPARATIVE APPRAISAL OF PHYSICAL, CHEMICAL, INSTRUMENTAL AND SENSORY EVALUATION METHODS FOR MONITORING OXIDATIVE DETERIORATION OF GHEE(AAU, Anand, 2014) MEHTA, BHAVBHUTI MANOJBHAI; Aparnathi, K. D.Ghee is the most famous traditional dairy product in India, now also gaining popularity in Asian countries as well as other continents like Australia, Europe and America. This fat rich dairy product is highly regarded for typical pleasing flavor. Ghee, being a lipid, prone to oxidation. Lipid oxidation is a degradation process considered to be a major cause of deterioration in the quality of fat products. It imparts rancid and unpleasant flavors to the products and thus decreases their organoleptic value. The reaction of fat oxidation is autocatalytic and occurs in two stages. In primary stage of the fat oxidation reaction between the unsaturated fatty acid and oxygen leads to formation of hydroperoxides and during secondary stage of the reaction the hydroperoxides decomposed into off smelling volatile compounds. This is very complex process that resuh in the formation of several different compounds. Therefore, there is no single method to measure all these compounds, consequently, the use of more than one testing method is recommended to monitor oxidative changes taking place in lipid. Numerous physical, chemical and instrumental methods have been reported to measure oxidative deterioration of oils and fats. The selection of appropriate analytical method is very essential to obtain correct information about oxidative deterioration of product like ghee. Analytical methods most widely reported are weight gain, conjugated dienes, Kreis number, iodine value, free fatty acids content, peroxide value, TBA value, p- Anisidine value, TOTOX value, Carbonyl value and Rancimat. However, no systematic work has been carried out so far for selection of the most appropriate methods to monitor oxidative deterioration of ghee. Therefore, this study was undertaken with a goal to compare performance of selected physical, chemical and instrumental methods to evaluate oxidative deterioration of ghee and to establish correlation between results obtained by these methods with results obtained by sensory evaluation of the ghee for flavor score. The entire work of study was divided as (1) selection of methods used for determination of peroxide value of ghee, (2) evaluation of methods used for monitoring oxidation of ghee on the bases of changes taking place during primary stage of lipid oxidation, (3) examination of methods used for monitoring oxidation of ghee on the bases of changes taking place during secondary stage of lipid oxidation, (4) testing of Rancimat for suitability to study oxidation of ghee, (5) comparison between performance of the method selected from stage of primary oxidation, method selected from secondary stage of oxidation and Rancimat to study oxidative deterioration of ghee and (6) work out the correlation between the results obtained by accelerated storage study and the results obtained from sensory evaluation of ghee for flavor score on storage at normal temperature. In selection of methods for determination of chemical changes taking place during primary stage as well as secondary stage of oxidation in ghee, accelerated storage study was employed. The samples of ghee were prepared by creamy butter method stored at 80°±2°C and monitor for chemical changes using selected methods at regular interval of every 48 hours. The samples of ghee were simultaneously analyzed for changes in flavor score by sensory evaluation using 9 point hedonic scale. For testing the suitability of Rancimat to study the oxidative deterioration of ghee, oxidation curves for the ghee were obtained at 90°C, 100°C, 110°C and 120°C and Arrhenius plots were obtained to predict the shelf life of ghee. Simultaneously ghee samples were also stored at 35°±2°C and monitor for changes in flavor score by sensory evaluation using 9 point hedonic scale at an interval of 10 days. Amongst the BIS, AOAC, AOCS/IUPAC, FOX and IDF methods used to monitor the peroxide formation in ghee, the FOX method was found most consistent. Moreover, the best correlation between changes in flavor score of ghee and peroxide formation was given by FOX method. Amongst the weight gain. Conjugated dienes, Kreis number. Iodine value, free fatty acids content and Peroxide value by FOX method based on chemical changes in the primary stage of the oxidation of ghee, except the FOX method no other method gave significant correlation with changes in flavor score of ghee during storage. Amongst the TBA value, p-Anisidine value, TOTOX value and Carbonyl value based on chemical changes in the secondary stage of the oxidation of ghee, the carbonyl value was found consistent. In addition it gave the highest correlation with changes in flavor score of ghee during storage. The changes in temperatures of Rancimat were highly correlated with induction period of ghee samples and changes in its flavor scores on storage at 80°±2°C. The predicted shelf life of ghee at 80°±2°C obtained from the prediction models prepared using the results of peroxide value by FOX method and carbonyl value of the ghee stored at 80°±2°C and that obtained from Rancimat was almost in corroboration with the actual shelf life of ghee stored at 80°±2°C. The predicted shelf life of ghee at 35°±2°C obtained from the prediction models prepared using the results of peroxide value by FOX method, carbonyl value and flavor score of the ghee stored at 80°±2°C was almost in corroboration with the actual shelf life of ghee stored at 35°±2°C. However, the predicted shelf life of ghee at 35°±2°C obtained from Rancimat was 2.47 times higher than the actual shelf life of ghee stored at 35°±2°C. Therefore, it appeared from the results that use of peroxide value by FOX method, carbonyl value and flavor score of ghee on storage at 80°±2°C are promising for predicting shelf life of ghee at normal temperature, but use of Rancimat does not appear promising to predict the shelf life of ghee on storage at normal temperature.ThesisItem Open Access BIOCHEMICAL AND MOLECULAR CHARACTERIZATION OF COLD-INDUCED SWEETENING IN POTATO (SOLANUM TUBEROSUM L.) VARIETIES DURING STORAGE(AAU, Anand, 2014) GALANI, YAMDEU JOSEPH HUBERT; TALATI, J. G.Potato (Solatium tuberosum L.) is the third most important food crop, the most important non-grain food crop and one of the most essential basic vegetable worldwide as well as in Indian subcontinent. After harvest, potatoes are stored in cold storage to provide round the year supply to markets and consumers. But during storage at cold temperatures, many cultivars accumulate free reducing sugars derived from breakdown of starch to sucrose that is ultimately cleaved by acid invertase to produce glucose and fructose in a metabolic process known as cold-induced sweetening (CIS). Understanding the basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars adversely affect the quality of processed food products. Investigations were carried out to characterize at biochemical and molecular levels the CIS in 11 potato varieties namely DSP 287, DSP 186, Kufri Surya, K. Chipsona-3, K. Sutlej, K. Sadabahar, K. Jyoti, K. Lauvkar, K. Himsona, K. Bahar and K. Badshah stored at 3 different temperatures viz., room temperature (25-32°C), incubator (15°C) and cold storage (4°C). Samples, collected every 15 days intervals for 105 days storage were analyzed for different parameters associated with carbohydrate metabolism on one side, and antioxidant capacity on the other side. Analysis of carbohydrate dynamics showed that low temperature storage negligibly influenced dry matter, starch and maltose contents of tubers, but a significant increase in reducing sugars, total soluble sugars, fructose, glucose, hexoses:sucrose ratio and a decrease of sucrose content were observed at 4°C as compared to room temperature. A strong positive correlation was found between reducing sugars and total soluble sugars, and between fructose and glucose. Additionally, important shrinkage and sprouting of tubers was observed at 15°C, they were less intense at room temperature, and no any shrinkage and sprouting occurred on tubers stored at 4°C. The potato varieties also appeared to be suitable for processing immediately after harvest or short storage at room temperature. The activity of β-amylase was considerably increased by storage at low temperature, and a weak correlation with starch content indicated an important role of other enzymes in starch degradation while absence of maltose accumulation with increased β-amylase activity implied a possible significant activity of maltase in potato tubers. Acid invertase activity drastically rose at low temperature and strongly paralleled reducing sugars, glucose, fructose and hexoses:sucrose ratio. Moreover, as acid invertase activity increased, sucrose content decreased, indicating the essential role of acid invertase in development of CIS. The above findings allowed to group the 11 potato varieties into low to high sugar-forming groups and thereby select K. Jyoti as CIS-tolerant and K. Badshah as CIS-susceptible. Banding pattern of both native and denaturated proteins of potato tubers could not clearly discriminate the varieties. Zymograms of p-amylase revealed differentially induced bands at low storage temperatures, which might justify the observed increase of enzyme activity. Screening of the 11 potato varieties with 10 SSR primers detected a total of 42 alleles arranged in 44 different configurations, among which 37 alleles (88%) were polymorphic. The polymorphic information content value of the SSR locus ranged from 0.473 to 0.787 thus indicating a high utility of these markers for study of genetic diversity in potato. The dendrogram derived from Dice's similarity coefficients among the 11 varieties could partially but efficiently differentiate close parents and sugar-forming groups. Differential gene expression analysis showed that during storage expression of vacuolar acid invertase gene StvacINV1 and p-amylase gene BAMl increased at low temperature and their transcripts were more expressed in the CIS-tolerant variety than the CIS-sensitive. Expression of invertase inhibitor gene INH2a however was higher in the CIS-tolerant variety than the CIS-sensitive. Correlating StvacINVl and INH2a expressions with reducing sugar content and acid invertase activity established that post-translational regulation of acid invertase by the invertase inhibitor protein could be an important component of resistance to CIS. Besides, correlation between BAM1 expression and (β-amylase activity affirmed the hypothesis of several enzymes and pathways involved in starch degradation during cold storage of potato. Analysis of antioxidant capacity parameters revealed that low temperature storage greatly influenced vitamin C content as well as the phenolic content. During storage, both parameters initially increased, then a fluctuated decline was observed but until the last day of observation, they remained above the initial level. Phenolic acids profiling by UPLC identified 12 compounds among which the most abundant was chlorogenic acid followed by gallic acid, sinapic acid and ellagic acid which is reported for the first time, while trans-cinnamic acid was the lowest. Except paracoumaric acid which decreased at 4°C, all the phenolic acids increased with storage among which sinapic acid and feruhc acid appeared to be most enhanced. Correlation analysis showed that gallic acid, caffeic acid. chlorogenic acid and protocatechuic acid significantly contributed to total phenolic content. Evaluation of antioxidant activity showed a close relationship between DPPH and ABTS methods. Antioxidant activity estimated by both the methods increased up to 60 days storage then at 90 days, they dropped to a level comparable or lower than the original value, irrespective of the storage temperature. Correlation study revealed that chlorogenic acid, gallic acid and ferulic acid mostly contributed to antioxidant activity. Activity of the antioxidant enzymes superoxide dismutase and ascorbate peroxidase both increased initially but then decreased to values lower than the initial level and were not influenced by storage temperature. Correlation with antioxidant activity indicated that the enhancement of reactive oxygen scavenging species in tubers could result mainly from ascorbate peroxidase activity. Isoforms of the two enzymes showed interesting polymorphism and changes in bands intensity as well as differential induction or suppression of bands during storage. However, isozymes of ascorbate peroxidase showed higher similarity and better discrimination of the varieties. Although a clear relationship between CIS and antioxidant capacity was not established, nevertheless it appeared that low sugar-forming varieties K. Jyoti, K. Himsona and K. Surya were also having high antioxidant capacity whereas K. Chipsona-3 and K. Bahar both high sugarforming had low antioxidant capacity. Hence, it is not unreasonable to suggest that antioxidant capacity of potato tubers should be taken into account in development of CIS-resistant varieties. Nonetheless, additional evidences are needed to confirm this suggestion as well as there is an urgent need to develop new varieties capable to cope with this cold stress.