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ThesisItem Open Access Molecular characterization of indigenous chicken breeds using RAPD-PCR(Sher-e-Kashmir University of Agricultural Sciences & Technology, Jammu (J&K), 2023-03-27) Dixsha Bala; Nishant KumarThe present study was carried out on a total of hundred chicken including 50 each of Kadaknath and Aseel chicken breeds with the objectives to study genetic variation between two breeds of chicken using RAPD-PCR technique, to establish genetic relationship and genetic distance between these breeds. The blood samples of Kadaknath and Aseel chicken were collected from local farms of RS Pura, Jammu. DNA was isolated by HiPurA Blood Genomic DNA Miniprep Purification Kit (Hi-media). Six primers were screened for RAPD study of two chicken breeds. But, only four primers generated reproducible and distinct RAPD profiles. A total of 15 loci were found in Kadaknath and Aseel chicken by four different RAPD primers out of which 9 were polymorphic. Average no. of bands ranged from 1.66 to 4.94. The mean effective number (ne) of allele for Kadaknath was 1.5886 ± 0.4977 and Nei’s gene diversity (h) was 0.2971 ± 0.2511. The Shannon’s information index (I) was 0.4130 ± 0.3491 in Kadaknath. The mean effective number (ne) of allele for Aseel was 1.5642 ± 0.4809 and Nei’s gene diversity (h) was 0.2902 ± 0.2459. The Shannon’s information index (I) was 0.4060 ± 0.3436 in Aseel. The genetic identity between Kadaknath and Aseel breeds was high and it was 0.9886. On the other hand the genetic distance between Kadaknath and Aseel was 0.0114. The genetic diversity reported among chicken breeds should be expanded by exploring more advanced analysis of genetic markers.ThesisItem Open Access Molecular Characterization by Microsatellite Markers and Immune Related Genes in Poonchi Chicken(Sher-e-Kashmir University of Agricultural Sciences & Technology-Jammu (J&K), 2022-11-09) Azad, Mandeep Singh; Chakraborty, DibyenduIndigenous / native population is very important source of genes for future breeding and improvement purposes. They are more adapted to the local environment and have a natural resistance to some diseases. Therefore, the present study was undertaken with the objectives to characterize the Poonchi chicken population by microsatellite markers and to characterize the immune related genes namely ChB6, IAP-1 and IL-15Rα. A total 50 unrelated blood samples of indigenous chicken from its natural breeding tract were utilized for molecular characterization. Genomic DNA was isolated. Ten (10) FAO recommended microsatellite markers namely ADL0268, MCW0206, MCW0081, ADL0278, MCW069, MCW0248, MCW0111, MCW0222, MCW0016, and LEI0094 were used for molecular characterization. PCR amplifications were performed with some modifications and PCR products were run in 10% Urea-PAGE for genotyping. Microsatellite data were analyzed by using molecular biology software Popgen32. For molecular characterization of immune related genes RNA was isolated and cDNA was synthesized. Primers were designed and PCR amplification was performed. Sequencing of the products was done and results were analyzed by using MEGAX and BioEdit softwares. A total of 72 alleles across 10 different microsatellite loci under present study were observed with mean number of loci as 7.200±1.2293. The Chi-square test revealed deviation from HWE. The mean na, ne and Shannon’s information index (I) in the population were 7.2000, 5.2928 and 1.7648, respectively. The PIC values for different microsatellite loci ranged from 0.6348 (LEI0094) to 0.9751 (MCW0016). The overall average inbreeding coefficient (FIS) was -0.0416. The results obtained from sequence analysis showed that there was no variation in the sequences of Poonchi, Kadaknath and Leghorn chicken populations for ChB6 and IAP-1 genes but there was variation with Fayoumi and other reported Gallus gallus sequences and other species. For IL-15Rα gene there was variation within Poonchi population as well as between different chicken populations like Kadaknath, Leghorn, Fayoumi. From the present study it can be recommended that further studies on large population should be carried out to explore the molecular level variations for formulation of breeding strategies, genetic improvement and conservation of indigenous chicken populationThesisItem Open Access Study on growth and morphometric traits of quail reared under sub-tropical region of Jammu(Sher-e-Kashmir University of Agricultural Sciences & Technology of Jammu (J&K), 2022-09-26) Parihar, Shimona; Kumar, DhirendraQuail farming is becoming an in-demand enterprise, due to lesser capital requirements and better profit margins. The study evaluated the effects of variety on growth and morphometric traits of two varieties (Pharaoh and Pearl variety) of quail. A total of 250 birds (125from each variety) and 300 eggs were observed and studied. Observations were recorded for various phenotypic traits (plumage colour, skin colour, shank colour and eye colour), morphometric traits (body length, body girth, wing length, shank length, shank diameter, drumstick length, breast width, pelvic width and keel length) and growth traits (body weight and body weight gain), along with egg quality traits. A total of 300 eggs (100 per month) were also procured and evaluated for egg quality parameters for three months. Data collected were analyzed by SPSS, SPAB and R studio soft-wares. Sexual dimorphism based on plumage colour was observed in Pharaoh variety and was absent in Pearl.The results showed that variety had impact on the linear body measurements at different age groups, except shank diameter and breast width. The effect of variety on body weight was also very significant at different ages. The age groups had highly significant effect on body weight gain in both the varieties. The effect of sex on body weight gain was also highly significant, with females being heavier than males in both the varieties. Both the varieties showed highest body weight gain during 3-4 week of age. The average values for Egg weight (11.35 g) and Haugh Unit (94.96%) showed highest values during third month. Pearl variety showed better performance for all the traits than Pharaoh variety. The present scientific study could be considered as pilot study which can provide a base line information for further studies and can also provide optimal information for those interested in quail farming in sub-tropical region of Jammu.ThesisItem Open Access Phenotypic characterization of local cattle population at Poonch and Rajouri districts in UT of Jammu and Kashmir(Sher-e-Kashmir University of Agricultural Sciences & Technology of Jammu (J&K), 2022-09-21) Farid, Asma; Kumar, DhirendraPhenotypic characterization of local cattle population is important to provide overall picture of cattle genetic diversity. Local cattle population is mainly reared by small and marginal farmers in UT J&K. The present study was conducted with the objectives to study the phenotypic, morphometric, production & reproductive traits of the local cattle in Poonch & Rajouri districts of J&K. A stratified random sample survey was conducted in different blocks namely Poonch, Mandi, Loran, Sathra and surankote in Poonch district and five blocks of district Rajouri namely Rajouri, Darhal, Thanamandi, Budhal & Plangar. Data of total 120 animals (32 male & 88 females) were collected randomly from these blocks. From each district data of total 60 cattle of different age and sex were recorded for the study. Four different body colors i.e, black (B), white (W), brown (Br) & roan (Rr) were found. Statured cattle were categorized as short & medium. The forehead of this local cattle was concave in majority of the animals. Ears were horizontally oriented. In most of cases tail was almost touching the ground with a prominent tail switch in both the sexes. Hump size in local cattle varied from small to medium size in cows and medium to large size in bulls. Cattle from Poonch region were better in production traits like lactation length (200±3.4) days, milk yield per day (kg) (2.4±0.1) and lactation yield (450±9.1) kg than cattle from Rajouri region having lactation length (174±4.8) days, milk yield per day (kg) (1.7±0.08) and lactation yield (381±8.5) kg. Also, it was found that sex and age group (AG) had highly significant (P<0.01) effect on all morphometric traits i.e chest girth (CG), height at wither (HW) and body length (BL). However, geographical region had no significant effect on any of the morphometric traits. Further, concave forehead, long tail almost touching the ground & ears having horizontal orientation can be considered as distinguishing features of these cattle.ThesisItem Open Access Growth hormone (GH) 1 gene polymorphism and its association with growth traits in quail(Sher-e-Kashmir University of Agricultural Sciences & Technology of Jammu (J&K), 2022-09-22) Ahamed, Naseer; Ahamed, Naseer; Ahamed, Naseer; Mahajan, Vikas; Mahajan, Vikas; Mahajan, VikasThe aim of this study was to investigate polymorphism of growth hormone (GH) 1 gene and its association with growth traits in quail using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Blood samples of 100 quails along with their body weight at 0, 1, 2, 3, 4, 5 and 6 weeks of age were taken. Genomic DNA was extracted by using DNA kit (HIMEDIA). The amplified PCR product of GH1 gene was 776 bp in size and was digested with Msp1 restriction enzyme at 37 °C for 30 min. The resulted digested products were run on 2.5% agarose gel, showed three different genotypes: AA (539 bp, 237 bp), AB (776 bp, 539 bp, and 237 bp) and BB (776 bp). The genotypic frequencies were found to be 0.50 for the AA genotype, 0.43 for the AB genotype and 0.07 for the BB genotype, while the gene frequencies for A and B allele were 0.715 and 0.285 respectively. The population was in Hardy-Weinberg equilibrium (HWE) as evidenced by the non-significant Chi-square value. The polymorphic information content value showed moderate level of polymorphism of 0.32 that indicates the effectiveness of the marker. Least square analysis of variance revealed non-significant differences at 0, 1, 2, 3, and 4 weeks of age with respect to different genotypes of GH1 gene, whereas highly significant (P<0.01) differences were observed at 5th and 6th week of age. BB genotype has significantly (P<0.01) higher body weight followed by AB and AA genotype at 5th and 6th week of age. The results indicate that the GH1 gene may be used as a candidate gene to improve the growth traits and also selection of superior bird can be done at any early age of 5 weeks for the genetic improvement in the quail.ThesisItem Open Access Molecular characterization of HSP70 gene in local chicken of Poonch(Sher-e-Kashmir University of Agricultural Sciences & Technology, Jammu (J&K), 2022-02-02) Singh, Harpreet; Kumar, NishantHeat shock protein 70 (HSP70) plays a variety of functions in the cells and is responsible for cytoprotection under stress conditions. In this study, molecular characterization of HSP70 gene in local chicken of Poonch was undertaken with the objectives of amplifying and sequencing of HSP70 gene and studying the genetic similarity and distance between different species with respect to local chicken of Poonch. Total RNA was extracted from freshly collected chicken blood and cDNA was synthesized by reverse transcription. HSP70 gene was amplified using specific primers designed by Primer 3 software. The amplified product (1986 bp) was purified and ligated in pGEM T Easy vector. The ligated product was cloned using DH5α competent cells and then sequenced. Sequence analysis of the HSP70 gene was done using MEGA X and DNA STAR softwares. The comparison of HSP70 CDS sequence demonstrated 100%, 95.96%, 95.85%, 95.70%, 92.60%, 91.44% 91.39%, 91.39%, 81.47%, 81.41% and 81.36% similarity with chicken, quail, turkey, guinea fowl, pigeon, ostrich, emu, duck, goat, sheep and pig, respectively indicating a high sequence homology among different species. The percentage identity of translated protein sequence of 634 residues of HSP70 gene was 100%, 98.26%, 98.11%, 98.11%, 97.95%, 97.95%, 97.48%, 97.32%,92.36%, 91.82% and 91.51% similar to chicken, quail, turkey, guinea fowl, pigeon, duck, emu, ostrich, pig, sheep and goat, respectively. The phylogenetic tree drawn by MEGA X software showed that the gene is evolutionarily conserved. Z test was conducted which showed that the gene might have evolved by purifying selection (dS>dN).ThesisItem Open Access Molecular characterization of HSP60 gene in Poonchi chicken(Sher-e-Kashmir University of Agricultural Sciences & Technology, Jammu (J&K), 2022-01-17) Kour, Komal Deep; Chakraborty, DibyenduHeat shock protein 60 (HSP60) is a multifunctional protein, playing a crucial role in protecting organisms from environmental stress. The present study molecular characterization of HSP60 gene in Poonchi chicken was undertaken with the objectives of amplifying and sequencing of HSP60 gene and studying the genetic similarity and distance between different species to local chicken of Poonch. Total RNA was extracted from freshly collected chicken blood and cDNA was synthesized by reverse transcription. HSP60 gene was amplified from cDNA using specific primers designed by Primer 3 software. The amplified product was purified and thereafter cloned and sequenced. A partial cDNA sequence of chicken HSP60 gene of 1002bp encoding 327 amino acids was obtained. On its comparison, chicken partial cDNA sequence demonstrated 91.30%, 90.61%, 83.63%, 97.46%, 96.85%, 95.83%, 82.67%, 92.35% and 93.37% homology with pigeon, ostrich, rabbit, guinea fowl, turkey, quail, guinea pig, geese and duck, respectively. Deduced amino acid sequence of 327 residues of chicken HSP60 gene was 98.47%, 96.94%, 96.64%, 99.30%, 98.17%, 99.00%, 96.33%, 98.47% and 99.08% similar to pigeon, ostrich, rabbit, guinea fowl, turkey, quail, guinea pig, geese and duck, respectively. The phylogenetic tree showed conserved nature of HSP60 gene. Maximum divergence and minimum divergence were observed with guinea pig CDS and guinea fowl, respectively. In Z test it was found selection is of purifying type. It can be concluded from the present study that chicken HSP60, is highly conserved among different species and HSP60 gene might have been evolved by purifying selection (dS > dN).ThesisItem Open Access Molecular characterization of HSP108 gene in local chicken of Poonch(Sher-e-Kashmir University of Agricultural Sciences & Technology, Jammu (J&K), 2022-01-17) Singh, Vishav Jeet; Mahajan, VikasThe present study on molecular characterization of heat shock protein 108 (HSP108) gene in local chicken of Poonch was undertaken with the objective of amplification and sequencing of HSP108 gene and to study the genetic similarity as well as distance between different species. Total RNA was extracted from 2ml freshly collected chicken blood using all blood RNA purification kit. cDNA was synthesized by reverse transcription from RNA using RevertAid First Strand cDNA Synthesis kit. HSP108 gene was amplified from cDNA using specific primers designed by Primer3 software. The amplified product was purified by GenElute gel extraction kit and thereafter cloned and sequenced. Coding sequence (CDS) of HSP108 gene was 2388 bp that encoding 795 amino acids. On its comparison, chicken CDS demonstrated 97.24, 96.94, 96.61, 93.94, 83.23, 83.13, 82.29, 82.85, 82.76, 82.49 and 82.44 percent homology with turkey, quail, guinea fowl, duck, horse, camel, pigs, cattle, buffalo, sheep and goat, respectively. Indicates close evolutionary relationship and high sequence homology among the species. Deduced amino acid sequence of 795 residues of HSP108 gene was 98.82, 97.51, 52.13, 51.49, 51.15, 48.22, 47.86, 47.86, 47.86, 47.86, 47.66 and 47.62 percent similar to chicken, duck, pig, buffalo, camel, goat, quail, turkey, sheep, horse, cattle and guinea fowl, respectively. Phylogenetic analysis revealed that chicken CDS sequence was close to turkey, followed by quail, guinea fowl and duck but it was most divergent from equines. It may also be inferred that this gene might have been evolved by purifying selection. From the present study, it can be concluded that chicken HSP108 is highly conserved among different species.ThesisItem Open Access Molecular characterization of HSP70-1 gene in Beetal goat(Sher-e-Kashmir University of Agricultural Sciences & Technology, Jammu (J&K), 2021-04-15) Khan, Kashif Dawood; Kumar, Dr. NishantHSP70 play a variety of functions in the cells and are responsible for cytoprotection under stress conditions. The present study was conducted to carry out molecular characterization of HSP70-1 gene in Beetal goat and analyzing genetic diversity through sequences available in the databases. Genomic DNA was extracted from whole blood of Beetal goat. HSP70-1 gene was amplified using specific primers designed by Primer 3 software. The amplified PCR product (1917 bp) was sequenced by Sangers dideoxy chain termination method with “Primer Walking”. A partial cDNA sequence of goat HSP70-1 gene of 1839 bp encoding 613 amino acids was obtained. Comparison of nucleotide and deduced amino acid sequences of Beetal goat HSP70-1 gene was done with different species of animals. The comparison of Beetal goat HSP70-1 partial cDNA sequence demonstrated 99.08, 98.10 and 97.66 percent homology with sheep, buffalo and cattle, respectively which indicates close evolutionary relationship and high sequence homology among the species. Deduced amino acid sequence of 613 residues of Beetal goat HSP70-1 gene was 98.83 and 98.99 percent similar to cattle and buffalo, respectively. The phylogenetic tree drawn by MEGA X software at nucleotide level showed conserved nature of HSP70-1 gene. The pair wise distance between sequences aligned with ClustalW method was estimated by MEGA X software. Maximum divergence from Beetal partial cDNA was observed with chicken CDS and minimum with sheep. Z test based on relative abundance of synonymous and non synonymous substitution indicates purifying selection and not positive selection operating on HSP70-1 gene.
