Molecular Characterization of Rathi and Tharparkar Indigenous Cattle (Bos indicus) Breeds by RAPD-PCR

Sharma, Amit Kumar,Bhushan, Bharat,Kumar, Sanjeev,Kumar, Pushpendra,Sharma, Arjava,Kumar, Satish Asian Australasian Association of Animal Productio 2004 Animal Bioscience Vol.17 No.9

Random amplification of polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR) analysis was carried out using DNA samples of 30 animals of Rathi cattle and 42 animals of Tharparkar cattle. Genomic DNA was isolated as per standard protocol and evaluated for its quality, purity and concentration. Twenty three random primers were screened out of which 15 primers yielded satisfactory amplifications and were used for further analysis. Average numbers of polymorphic fragments per primer were 7.07${\pm}$0.86 in Rathi and 6.80${\pm}$0.61 in Tharparkar cattle. The percentage of polymorphic bands in these two cattle breeds were 86 and 87%, respectively. Within breed genetic similarities for pooled over primers in the animals of Rathi and Tharparkar breeds were .577${\pm}$0.30 and 0.531${\pm}$0.02, respectively on the basis of band frequency (BF) and 0.645${\pm}$0.04 and 0.534${\pm}$0.04, respectively on the basis of band sharing (BS). Averages of between breed genetic similarities for pooled over primers were 0.97 and 0.92 according to BF and BS, respectively, which reflect higher degree of genetic similarity between Rathi and Tharparkar cattle breeds. Index of genetic distance based on BF and BS for pooled over primers was 0.030${\pm}$0.011 and 0.088${\pm}$0.031, respectively. Percentage of polymorphic bands and within-breed genetic similarities on the basis of band frequency (BF) and band sharing (BS) for pooled over primers revealed higher genetic similarity in Rathi than Tharparkar cattle population. High estimates of between breed genetic similarities for pooled over primers indicated that either Rathi is having decent from Tharparkar or both the cattle breeds are having common descent. Low value of Index of genetic distances between these two cattle breeds may be due to the fact that Rathi and Tharparkar cattle breeds are the native of Thar Desert in Northwest India. The results of between breed genetic distances also confirm the existence of high degree of genetic similarity between these two breeds of cattle.

Multi-objective optimization of electric-discharge machining process using controlled elitist NSGA-II

Pushpendra S. Bharti,S. Maheshwari,C. Sharma 대한기계학회 2012 JOURNAL OF MECHANICAL SCIENCE AND TECHNOLOGY Vol.26 No.6

Parametric optimization of electric discharge machining (EDM) process is a multi-objective optimization task. In general, no single combination of input parameters can provide the best cutting speed and the best surface finish simultaneously. Genetic algorithm has been proven as one of the most popular multi-objective optimization techniques for the parametric optimization of EDM process. In this work, controlled elitist non-dominated sorting genetic algorithm has been used to optimize the process. Experiments have been carried out on die-sinking EDM by taking Inconel 718 as work piece and copper as tool electrode. Artificial neural network (ANN) with back propagation algorithm has been used to model EDM process. ANN has been trained with the experimental data set. Controlled elitist non-dominated sorting genetic algorithm has been employed in the trained network and a set of pareto-optimal solutions is obtained.

Comparative analysis of VMT genes/proteins in selected plant species with emphasis on bread wheat (Triticum aestivum L.)

Sharma Hemant,Shayaba,Kumar Rahul,Kumar Jitendra,Bhadana Deepa,Batra Ritu,Singh Rakhi,Kumar Sachin,Roy Joy K,Balyan Harindra S.,Gupta Pushpendra K. 한국유전학회 2023 Genes & Genomics Vol.45 No.11

Background In recent years, the study of molecular basis of uptake, transport and utilization of grain Fe/Zn (GFe/GZn) in wheat has been an active area of research. As a result, it has been shown that a number of transporters are involved in uptake and transport of Fe. In a recent study, knockout of a transporter gene OsVMT (VACUOLAR MUGINEIC ACID TRANSPORTER) in rice was shown to be involved in Fe homoeostasis. Objective In this study, we analysed VMT genes among six monocots and three dicots with major emphasis on wheat VMT genes (TaVMTs), taking OsVMT gene as a reference. Methods and results Using OsVMT gene as a reference, VMT genes were identified and sequence similarities were examined among six monocots and three dicots. Each VMT protein carried one functional domain and 7 to 10 distinct motifs (including 9 novel motifs). The qRT-PCR analysis showed differential expression by all the six TaVMT genes in pairs of contrasting wheat genotypes with high (FAR4 and WB02) and low (K8027 and HD3226) GFe/GZn at two different grain filling stages (14 DAA and 28 DAA). TaVMT1 genes showed up-regulation in high GFe/Zn genotypes relative to low GFe/Zn genotypes, whereas the TaVMT2 genes showed down-regulation or nonsignificant up-regulation in a few cases. Conclusions At 14 DAA, each of the six TaVMT genes exhibited higher expression in wheat genotypes with high GFe and GZn relative to those with low GFe and GZn, suggesting major role of VMT genes in improvement of grain Fe/Zn homoeostasis, thus making TaVMT genes useful for improvement in Fe/Zn in wheat grains.

PCR-SSCP of Serum Lysozyme Gene (Exon-III) in Riverine Buffalo and Its Association with Lysozyme Activity and Somatic Cell Count

Sahoo, Nihar Ranjan,Kumar, Pushpendra,Bhushan, Bharat,Bhattacharya, T.K.,Sharma, Arjava,Dayal, Sanker,Pankaj, Prabhat Kumar,Sahoo, Monalisa Asian Australasian Association of Animal Productio 2010 Animal Bioscience Vol.23 No.8

Serum lysozyme gene is one of the important genes influencing the immune system as its product can cause lysis of bacterial cell wall by cleaving the peptidoglycan layer. The present investigation on the serum lysozyme gene of Indian riverine buffalo was undertaken with the objectives to identify and characterize single nucleotide polymorphic patterns by PCR-SSCP method as well as to study the effect of different genotypes on serum lysozyme activity and somatic cell count. A total of 280 animals comprising four different famous bubaline breeds (Murrah, Mehsana, Surti and Bhadawari), spread over six different farms across the country were used for this study. A 276 bp (partial intron 2, complete exon 3 and partial intron 3) fragment of lysozyme gene was screened for polymorphism using the SSCP technique. Four genotypes namely AA, AB, BC and AC were observed, out of which BC genotype was found to be the most frequent. Among these three alleles, C allele (0.38) was most prevalent in these populations. Various SSCP allelic variants were cloned for sequencing and sequences were submitted to NCBI Genbank. From the alignment of the nucleotide sequences of various allelic variants, it was found that there were differences in 12 positions among the alleles, out of which maximum variation (at 8 places) was found in the intronic region. The allele A was closer to allele-C than allele-B. Allele B was phylogenetically equidistant from both of the other alleles. Mean lysozyme activity determined in serum samples of different animals of Murrah buffalo was $27.35{\pm}2.42\;{\mu}g$ per ml of serum, whereas the mean somatic cell count was $1.25{\pm}0.13{\times}10^5$ cells per ml of milk. The SSCP pattern-wise effects of various genotypes on lysozyme activity and SCC were analyzed. Although the mean values were apparently different in various genotypes, these differences were statistically non-significant. It can be concluded that the riverine buffaloes are sufficiently polymorphic with respect to serum lysozyme gene. The absence of AA genotype in Bhadawari breed of buffalo can be considered as a marker for breed characterization. The difference of four nucleotides in exon-3 indicates high selection pressure on the gene.

Actinobacteria from Cow Feces: Isolation, Identification and Screening for Industrially Important Secondary Metabolites

( Preeti Semwal ),( Vinay Rawat ),( Pushpendra Sharma ),( Mamta Baunthiyal ) 한국미생물생명공학회(구 한국산업미생물학회) 2018 한국미생물·생명공학회지 Vol.46 No.1

Actinobacterial strains isolated from Cow feces were studied for their antifungal attributes against phytopathogens and industrially important enzymes. A total of 30 Actinobacterial strains were obtained from 10 samples of cow feces. All the strains were belonging to the genera Streptomyces on the basis of morphological and chemotaxonomic analysis. During preliminary screening, out of 30 strains, 15 strains (50%) showed antifungal activity against five fungal phytopathogens including Aspergillus niger, Fusarium solani, Fusarium oxysporum, Macrophomina phaseolina and Rhizoctonia solani. While, isolate GBTCF-26 was found to be most active against R. solani with 62.2% inhibition of fungal mycelium, GBTCF-09 was prominent against F. solani and F. oxysporum with percent inhibition of 61.1% and 58.8%, respectively. Out of 30 strains, 19 (63.3%), 16 (53.3%), 11 (36.7%), 10 (33.3%), 4 (13.3%) and 8 (26.7%) strains were producing amylase, caseinase, gelatinase, lipase, chitinase and cellulose, respectively. The selected strains, GBTCF-09, GBTCF- 21 and GBTCF-26, were identified as Streptomyces sp. on the basis of their 16S rDNA sequence. The study supports the idea that the Actinobacteria from unique niches (Cow feces) possess the production potential of industrially important enzymes including bioactive molecules.

Nucleotide Sequencing and PCR-RFLP of Insulin-like Growth Factor Binding Protein-3 Gene in Riverine Buffalo (Bubalus bubalis)

Padma, B.,Kumar, Pushpendra,Choudhary, V.,Dhara, S.K.,Mishra, A.,Bhattacharya, T.K.,Bhushan, B.,Sharma, Arjava Asian Australasian Association of Animal Productio 2004 Animal Bioscience Vol.17 No.7

Insulin-like growth factor binding protein-3 (IGFBP-3) gene is a structural gene associated with the growth and development of the animals. The present investigation was carried out to unravel nucleotide sequence and polymerase chain reactionrestriction fragment polymorphism (PCR-RFLP) of IGFBP-3 gene in buffalo. Genomic DNA was isolated from a total of 157 animals belonging to Murrah, Surti, Jaffarabadi and Nagpuri breeds of Indian riverine buffalo. A 655 bp of IGFBP-3 gene was amplified in all the breeds and amplicons were digested with Hae III, Taq I and Msp I restriction enzymes. On digestion with Hae III yielded single restriction pattern of 8 fragments of sizes 201, 165, 154, 56, 36, 19, 16 and 8 bp in all the animals studied. Similarly Taq I and Msp I also revealed single restriction pattern yielding fragments of sizes 240 and 415 bp and 145 and 510 bp, respectively. This shows nonpolymorphic nature of restriction sites in buffalo. Nucleotide sequencing of 587 bp of IGFBP-3 gene in Murrah buffalo was done and submitted to the GenBank (Accession No. AY304829). Nucleotide sequencing revealed an addition of 4 bases in the intronic region as compared to cattle.

Actinobacteria from Cow Feces: Isolation, Identification and Screening for Industrially Important Secondary Metabolites

Semwal, Preeti,Rawat, Vinay,Sharma, Pushpendra,Baunthiyal, Mamta The Korean Society for Microbiology and Biotechnol 2018 한국미생물·생명공학회지 Vol.46 No.1

Actinobacterial strains isolated from Cow feces were studied for their antifungal attributes against phytopathogens and industrially important enzymes. A total of 30 Actinobacterial strains were obtained from 10 samples of cow feces. All the strains were belonging to the genera Streptomyces on the basis of morphological and chemotaxonomic analysis. During preliminary screening, out of 30 strains, 15 strains (50%) showed antifungal activity against five fungal phytopathogens including Aspergillus niger, Fusarium solani, Fusarium oxysporum, Macrophomina phaseolina and Rhizoctonia solani. While, isolate GBTCF-26 was found to be most active against R. solani with 62.2% inhibition of fungal mycelium, GBTCF-09 was prominent against F. solani and F. oxysporum with percent inhibition of 61.1% and 58.8%, respectively. Out of 30 strains, 19 (63.3%), 16 (53.3%), 11 (36.7%), 10 (33.3%), 4 (13.3%) and 8 (26.7%) strains were producing amylase, caseinase, gelatinase, lipase, chitinase and cellulose, respectively. The selected strains, GBTCF-09, GBTCF-21 and GBTCF-26, were identified as Streptomyces sp. on the basis of their 16S rDNA sequence. The study supports the idea that the Actinobacteria from unique niches (Cow feces) possess the production potential of industrially important enzymes including bioactive molecules.

Effect of Leptin and IGFBP-3 Gene Polymorphisms on Serum IgG Level of Cattle Calves

Choudhary, Vivek,Kumar, Pushpendra,Saxena, V.K.,Bhattacharya, T.K.,Bhushan, Bharat,Sharma, Arjava,Ahmed, K.A. Asian Australasian Association of Animal Productio 2006 Animal Bioscience Vol.19 No.8

Leptin and IGFBP-3 are two proteins that play an important role in growth and metabolism of the animals. They are also involved in the immune function of animals and, thus, are candidate genes for the study of association with immune functions. Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) of these two genes was done to screen 64 crossbred (Holstein Friesian${\times}$Hariana) female calves of one year of age. From each RFLPs (fragments) three genotypes were observed. In all the RFLPs the mutant homozygotes were very less in numbers and, hence, were excluded from the least squares analysis. The serum IgG level was estimated using SRID assay. The mean level of serum IgG was $28.83{\pm}2.73mg/ml$. The effect of these identified genotypes on serum IgG level of calves at one year of age was analysed using least squares analysis. The HaeIII RFLP-AB genotype had significantly (p<0.05) higher serum IgG level ($31.86{\pm}3.05$) than the HaeIII RFLP-AA ($25.62{\pm}2.96$) genotype. There was no significant effect of leptin genotypes on the IgG level. The present results indicated a role of the IGFBP-3 gene on serum IgG level of cattle calves.

Effect of Butyrophilin Gene Polymorphism on Milk Quality Traits in Crossbred Cattle

Bhattacharya, T.K.,Misra, S.S.,Sheikh, Feroz D.,Sukla, Soumi,Kumar, Pushpendra,Sharma, Arjava Asian Australasian Association of Animal Productio 2006 Animal Bioscience Vol.19 No.7

A genetic polymorphism study on butyrophilin gene was carried out to explore variability of this gene and to estimate effects of such variability on milk quality traits in crossbred cattle. Polymorphism was unraveled by conducting Hae III PCR-RFLP of this gene. Three genotypes such as AA, BB and AB and two alleles namely A and B were observed in crossbred population. The frequencies of genotypes and alleles were 0.78, 0.17 and 0.04 for AA, AB and BB genotypes, respectively, and 0.87 and 0.13 for A and B alleles, respectively. The nucleotides, which have been substituted from allele A to B, were observed as C to G ($71^{st}$ nucleotide), C to T ($86^{th}$ nucleotide), A to T ($217^{th}$ nucleotide), G to A ($258^{th}$ nucleotide), A to C ($371^{st}$ nucleotide) and C to T ($377^{th}$ nucleotide). The nucleotide substitutions at $71^{st}$, $86^{th}$ and $377^{th}$ position of the fragment were found as silent mutations whereas nucleotide changes at $217^{th}$, $258^{th}$ and $371^{st}$ positions were detected as substitution of amino acid lysine with arginine, valine with isoleucine, and leucine with proline from allele A to B. The genotypes had significant effects ($p{\leq}0.05$) on total milk solid%, fat%, SNF%, while showing nonsignificant impact on total protein%. AA genotype produced highest average yield for all the traits.

Genetic Identity between Bhadawari and Murrah Breeds of Indian Buffaloes (Bubalus bubalis) Using RAPD-PCR

Saifi, H.W.,Bhushan, Bharat,Kumar, Sanjeev,Kumar, Pushpendra,Patra, B.N.,Sharma, Arjava Asian Australasian Association of Animal Productio 2004 Animal Bioscience Vol.17 No.5

Randomly Amplified Polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR) analysis was carried out with a battery of 11 random decamer primers to study band frequency (BF), genetic identity index (I) and mean average percentage difference (MAPD) between Bhadawari and Murrah breeds of buffalo. The primers OPA04 and BG15 resolved a band of 460 bp, which was present only in animals of Bhadawari breed. Whereas, the primers OPA14, BG27 and BG28 produced Murrah specific fragments of sizes 730 bp and 1,230 bp, respectively. The estimate of genetic identity index was highest (0.845) with the primer OPA01 and the lowest (0.479) with the primer BG27. The genetic identity index pooled over the primers was 0.596${\pm}$0.037 between these two breeds. The highest MAPD estimate (53.9) between the two breeds was obtained with the primer BG27 and the lowest (14.3) with the primer OPA01. It might be concluded that the genetic identity index between these two breeds calculated on the basis of BF showed moderate level of genetic identity with the primers employed. MAPD calculated on the basis of uncommon bands also demonstrated lower to medium level of genetic difference between Bhadawari and Murrah breeds of buffalo.