Elucidation of population stratifying markers and selective sweeps in crossbred Landlly pig population using genome-wide SNP data.

The present study was aimed at the identification of population stratifying markers from the commercial porcine SNP 60K array and elucidate the genome-wide selective sweeps in the crossbred Landlly pig population. Original genotyping data, generated on Landlly pigs, was merged in various combinations with global suid breeds that were grouped as exotic (global pig breeds excluding Indian and Chinese), Chinese (Chinese pig breeds only), and outgroup pig populations. Post quality control, the genome-wide SNPs were ranked for their stratifying power within each dataset in TRES (using three different criteria) and FIFS programs and top-ranked SNPs (0.5K, 1K, 2K, 3K, and 4K densities) were selected. PCA plots were used to assess the stratification power of low-density panels. Selective sweeps were elucidated in the Landlly population using intra- and inter-population haplotype statistics. Additionally, Tajima's D-statistics were calculated to determine the status of balancing selection in the Landlly population. PCA plots showed 0.5K marker density to effectively stratify Landlly from other pig populations. The A-score in DAPC program revealed the Delta statistic of marker selection to outperform other methods (informativeness and FST methods) and that 3000-marker density was suitable for stratification of Landlly animals from exotic pig populations. The results from selective sweep analysis revealed the Landlly population to be under selection for mammary (NAV2), reproductive efficiency (JMY, SERGEF, and MAP3K20), body conformation (FHIT, WNT2, ASRB, DMGDH, and BHMT), feed efficiency (CSRNP1 and ADRA1A), and immunity (U6, MYO3B, RBMS3, and FAM78B) traits. More than two methods suggested sweeps for immunity and feed efficiency traits, thus giving a strong indication for selection in this direction. The study is the first of its kind in Indian pig breeds with a comparison against global breeds. In conclusion, 500 markers were able to effectively stratify the breeds. Different traits under selective sweeps (natural or artificial selection) can be exploited for further improvement.

A Physiology-Based Mathematical Model to Understand Drug Delivery from Contact Lenses to the Back of the Eye.

OBJECTIVE: Therapeutic contact lenses, able to store drug and deliver it to the eye surface in a sustained fashion, gained interest as an effective and patient-friendly alternative to eye drops. Recent animal studies also demonstrated the presence of therapeutic drug levels in the back of the eye after wearing drug-loaded contact lenses, thus opening the possibility of treating the posterior segment without need of invasive intraocular injections. The drug pathways from contact lenses to the back of the eye require further investigation. METHODS: A mechanistic mathematical model was developed to evaluate the drug concentration over time in the tears, sclera and choroid, retina, aqueous humor and vitreous humor after the application of a therapeutic contact lens. The main drug transport mechanisms of the eye and the barrier properties of the different tissues were included in the model. Validation was performed by comparison with experimental data in literature. RESULTS: The model predictions of drug concentration over time reflected the experimental data both in the anterior and posterior segment of the eye. The model can differentiate between contributions to transport from different pathways. CONCLUSIONS: The model constitutes a first step towards the possibility of predicting the ocular drug distribution and the treatment efficacy in the early stage of contact lens development, and it may help reduce both the need for in vivo tests (with ethical and economic advantages) and the gap between the lens design and clinical application. It also allows for an improved understanding of drug transport in the eye.

Allele mining in gene encoding fc fragment of IgG receptor, transporter, alpha (FCGRT) and association of nucleotide variants with passive transfer of immunity in neonatal buffalo calves.

BACKGROUND: Neonatal Fc receptors (FcRn) mediate the transcytosis of IgG present in colostrum across absorptive gut epithelium of newborn calves. FcRn receptor is a heterodimer composed of two polypeptides encoded by FCGRT (Fc fragment of IgG Receptor Transporter neonatal) and B2M (Beta 2 microglobulin) genes. Polymorphism in FCGRT gene may have a bearing on absorption of colostral immunoglobulins by neonatal buffalo calves, thereby affecting their immune status and susceptibility to diseases. The primary aim of our study was to mine alleles and single nucleotide polymorphs in the FCGRT gene and determine their association with the levels of IgG in serum of neonatal buffalo calves. METHODS AND RESULTS: On the basis of serum IgG levels estimated by indirect ELISA in 80 newborn calves, 20 calves each with highest and lowest IgG concentration were selected to study polymorphism in the FCGRT gene. The exonic regions of this gene were amplified in nine fragments which were subjected to PCR-SSCP to detect variations followed by the sequencing of variants to locate the SNPs. A total of nine SNPs (7 in introns and 2 in exons) were detected in four polymorphic fragments. Association study based on Odds ratios (ORs) with 95% Confidence Interval (CIs) established that the SNP G40T in fragment 3 has a significant (P < 0.05) bearing on IgG level in serum of neonatal buffalo calves. CONCLUSION: Genetic variation in FCGRT gene in buffalo calves was found to be associated with their serum IgG levels in neonatal stage which may have implications in calf survival and growth vis-à-vis inadequate transfer of passive immunity.

Allelic to genome wide perspectives of swine genetic variation to litter size and its component traits.

Litter size is a complex and sex limited trait that depends on various biological, managemental and environmental factors. Owing to its low heritability it is inefficaciously selected by traditional methods. However, due to higher heritability of ovulation rate and embryo survival, selection based on component traits of litter size is advocated. QTL analysis and candidate gene approach are among the various supplementary/alternate strategies for selection of litter size. QTL analysis is aimed at identifying genomic regions affecting trait of interest significantly. Candidate gene approach necessitates identification of genes potentially affecting the trait. There are various genes that significantly affect litter size and its component traits viz. ESR, LEP, BF, IGFBP, RBP4, PRLR, CTNNAL1, WNT10B, TCF12, DAZ, and RNF4. These genes affect litter size in a complex interacting manner. Lately, genome wide association study (GWAS) have been utilized to unveil the genetic and biological background of litter traits, and elucidate the genes governing litter size. Favorable SNPs in these genes have been identified and offers a scope for inclusion in selection programs thereby increasing breeding efficiency and profit in pigs. The review provides a comprehensive coverage of investigations carried out globally to unravel the genetic variation in litter size and its component traits in pigs, both at allelic and genome wide level. It offers a current perspective on different strategies including the profiling of candidate genes, QTLs, and genome wide association studies as an aid to efficient selection for litter size and its component traits.

Genome wide copy number variations using Porcine 60K SNP Beadchip in Landlly pigs.

In the present study, Porcine 60K SNP genotype data from 69 Landlly pigs were used to explore Copy Number Variations (CNVs) across the autosomes. A total of 386 CNVs were identified using Hidden Markov Model (HMM) in PennCNV software, which were subsequently aggregated to 115 CNV regions (CNVRs). Among the total detected CNVRs, 58 gain, 49 were loss type while remaining 8 events were both gain and loss types. Identified CNVRs covered 12.5 Mb (0.55%) of Sus scrofa reference 11.1 genome. Comparison of our results with previous investigations on pigs revealed that approximately 75% CNVRs were novel, which may be due to differences in genetic background, environment and implementation of artificial selection in Landlly pigs. Functional annotation and pathway analysis showed the significant enrichment of 267 well-annotated Sus scrofa genes in CNVRs. These genes were involved in different biological functions like sensory perception, meat quality traits, back fat thickness and immunity. Additionally, KIT and FUT1 were two major genes detected on CNVR in our population. This investigation provided a comprehensive overview of CNV distribution in the Indian porcine genome for the first time, which may be useful for further investigating the association of important quantitative traits in Landlly pigs.Highlights115 CNVRs were identified in 69 Landlly pig population.Approximately 75% detected CNVRs were novel for Landlly population.Significant enrichment of 267 well-annotated Sus scrofa genes observed in these CNVRs.These genes were involved in different biological functions like sensory perception, meat quality traits, back fat thickness and immunity.Comprehensive CNV map in the Indian porcine genome developed for the first time.

Genetic polymorphism of microsatellite loci in MHC class II exon 2 gene and its association with endoparasitic infestation, predominantly Haemonchus contortus in Salem black goat.

MHC gene is highly polymorphic and plays central molecule in immune system in recognition of pathogens and parasites. The present study examined the variability of MHC class II DRB exon 2 gene using microsatellite analysis, and its association with Haemonchus contortus infestation in Salem Black goat population. Animals were naturally exposed to mixed infestation of endoparasites, predominantly Haemonchus contortus. Pooled fecal coproculture and larval identification showed predominant presence of haemonchus (L3) larva. Fecal egg count (FEC) and packed cell volume (PCV) were used as indicator traits. All the three studied loci, OarCP73, DYA, and ODRB1.2, were polymorphic having three, four, and four alleles and four, eight, and nine genotypes, respectively. The loci showed low to moderate values of polymorphic information content. The mean fecal egg count estimates were 477.12 ± 34.14 eggs per gram of feces and the mean PCV values were within the normal range; however, they showed negative correlation with FEC values. All the three microsatellite loci and the effect of genotypes on indicator traits were found to be statistically significant (p ≤ 0.05). The present study will be helpful in selection and breeding of goats for Haemonchosis resistance.

Estimation of microsatellite-based autozygosity and its correlation with pedigree inbreeding coefficient in crossbred cattle.

In countries where farming is largely subsistence, no pedigree records of farm animals are maintained at farmers' herd and scientific mating plans are not observed which leads to the accumulation of inbreeding and loss of production potential. Microsatellites have been widely used as reliable molecular markers to measure inbreeding. We attempted to correlate autozygosity estimated from microsatellite data with the inbreeding coefficient (F) calculated from pedigree data in Vrindavani crossbred cattle developed in India. The inbreeding coefficient was calculated from the pedigree of ninety-six Vrindavani cattle. Animals were further grouped into three groups viz. acceptable/low (F: 0-5%), moderate (F: 5-10%) and high (F: ≥10%), based on their inbreeding coefficients. The overall mean of the inbreeding coefficient was found to be 0.070 ± 0.007. A panel of twenty-five bovine-specific loci were chosen for the study according to ISAG/FAO. The mean FIS, FST, and FIT values were 0.0548 ± 0.025, 0.012 ± 0.001 and 0.0417 ± 0.025, respectively. There was no significant correlation between the FIS values obtained and the pedigree F values. The locus-wise individual autozygosity was estimated using the method-of-moments estimator (MME) formula for locus-specific autozygosity. The autozygosities ascribing to CSSM66 and TGLA53 were found to be significantly (p < .01 and p < .05, respectively) correlated with pedigree F values.

Genome-wide transcriptome profiling of CSF virus challenged monocyte-derived macrophages provides distinct insights into immune response of Landrace and indigenous Ghurrah pigs.

The present study was undertaken to characterize the distinct immune response in indigenous Ghurrah and exotic Landrace pigs by challenging monocyte-derived macrophages (MDMs) with CSF virus under in-vitro conditions and assessing the variations in the transcriptome profile at 48 h post-infection (hpi). RNA-sequencing was carried out in infected and non-infected MDMs of Ghurrah (n = 3) and Landrace (n = 3) piglets prior- as well as post-stimulation. MDMs of Ghurrah showed greater immune regulation in response to CSF infection with 518 significantly differentially expressed genes (DEG) in infected versus non-infected MDMs, as compared to only 31 DEGs in Landrace MDMs. In Landrace, the principal regulators of inflammation (IL1α, IL1ß and TNF) were upregulated in infected cells while in Ghurrah, these were downregulated. Overall, macrophages from indigenous Ghurrah showed more immunological dysregulation in response to virulent CSF virus infection as compared to the exotic Landrace pigs.

Impact of body condition on sow and litter performance, postpartum physiological, hematological, and biochemical parameters in Landlly crossbred pigs.

A total of 32 Landlly crossbred sows were categorized into three groups based on their body condition score (BCS) on the 90th day of gestation viz. low, moderate, and high body condition groups. BCS assessments were subsequently conducted on the 102nd day, at farrowing, and on the 7th, 17th, 27th, and 42nd day postpartum, along with measurements of ultrasonic backfat thickness and body weight. Key reproductive performance traits, including total number born (TNB) and alive (TLA), stillbirths, mummification, and litter weight at birth and weaning, were recorded. Several physiological, health, biochemical, and hematological parameters were also estimated at different lactation stages. Analysis of the data revealed that TNB, TLA (P<0.04), and weaned piglets (P<0.01), as well as litter weight at birth (P<0.08) and weaning (P<0.08), were significantly higher in the low BCS group. Meanwhile, the weaning to estrous interval (P<0.04) was optimized in the high and moderate BCS groups. However, sows with high body condition exhibited higher stillbirths and pre-weaning mortality. Additionally, the order of increasing body condition correlated with superior colostrum quality, characterized by higher IgG levels (P<0.02), and increased average milk yield during early and mid-lactation (P<0.03). A high occurrence of postpartum dysgalactia syndrome (PDS) was evident in obese sows (P<0.001). Moreover, a positive correlation was found between the body condition of sows at late gestation and cortisol concentration throughout lactation across the three BCS groups (P<0.0001). Furthermore, a strong positive correlation was observed between the postpartum serum concentration of T3 (thyroid hormone) (P<0.002) and the prepartum body condition of sows. Based on these findings, maintaining sows with a moderate body condition (BCS of 3) and a backfat thickness of 21 mm in the breeding herd is recommended for enhancing profitability and productivity at the farm level.

Detection of nucleotide variants in FCGRT (Fc fragment of IgG, receptor, transporter, alpha) gene and their influence on colostral IgG concentration in Indian water buffalo (Bubalus bubalis).

BACKGROUND: The neonatal Fc receptors (FcRn) mediate the transfer of immunoglobulin G (IgG) molecules from a dam's circulation to the colostrum produced by it immediately after parturition. In ruminants, the calves born are agammaglobulinemic therefore, ingestion of colostrum with high concentration of IgG imparts passive immunity to the newborn. The FcRn molecule is a heterodimer, coded by FCGRT (Fc fragment of IgG Receptor Transporter neonatal) and B2M (Beta 2 microglobulin) genes. Present study attempted to identify single nucleotide polymorphisms (SNPs) in the FCGRT gene in 40 buffaloes of Murrah breed and evaluated the association of these nucleotide variations and haplotypes with IgG concentration in their colostrum. METHODS AND RESULTS: Animals producing colostrum with high IgG and low IgG levels were identified by indirect ELISA and selected. SNPs were detected in the FCGRT gene sequence of selected animals by amplifying it in nine fragments covering all exons (with flanking introns) followed by PCR-single strand conformational polymorphism (PCR-SSCP). A total of nine SNPs were observed of which seven were present in flanking introns and two in exon 4 of the gene. The SNP A75G was non-synonymous and produced an amino acid change from isoleucine to valine. The exonic SNPs and corresponding haplotypes were found to be significantly (P < 0.01 and 0.05 respectively) associated with colostral IgG concentration based on Odds ratios at 95% confidence interval. CONCLUSION: Polymorphism in FCGRT gene is found to be associated with IgG concentration in colostrum and identification of females with desirable variations may prevent failure of passive transfer in neonatal ruminants.

Investigation of genetic polymorphism at ß-casein A1/A2 loci and association analysis with production & reproduction traits in Vrindavani crossbred cows.

Casein constitutes approximately 80% of the total protein in bovine milk and is regarded as a high-quality dietary protein embracing all the nine essential amino acids. However, the contested physiological effect of a bioactive peptide released upon digestion of a ß-casein milk protein variant originating from a cow of a particular genetic makeup has evoked wide interest in research and industry. Present investigations were carried out to genotype the polymorphism in milk ß-casein gene, delineate the seasonal, periodic, and parity variations in production and reproduction traits, and examine the genetic association between ß-casein genotypes and production, and reproduction traits in Vrindavani crossbred cows. The study revealed that all three types of genotypes viz. A1A1, A2A2 and A1A2 were present in the Vrindavani crossbred population with genotypic frequencies of 12.3%, 39.6% and 48.1% respectively. The least-squares analysis revealed that the season of calving, period of calving, and parity affected several production and reproduction traits of Vrindavani cows significantly. It was found that ß-Casein A1/A2 genotype had a significant effect on economic traits viz. LL (p ≤ 0.05), MY/LL (p ≤ 0.05), and Fat% (p ≤ 0.05) in Vrindavani crossbreds. The findings uncover the genetic constitution of the crossbreds for ß-casein locus and emphasize its relationship with important economic traits that can aid in devising selection goals.

Association of genetic variability in CD209 gene with bovine paratuberculosis disease: a case-control study in the Indian cattle population.

This study aimed to explore the association of single nucleotide polymorphisms (SNPs) in CD209 gene with the occurrence of bovine paratuberculosis (PTB) disease caused by Mycobacterium avium subspecies paratuberculosis (MAP) in Indian cattle. A total of 213 animals were preliminarily selected on the basis of physical body condition score, which was then screened by a panel of diagnostic tests viz. Johnin, ELISA, fecal microscopy, and fecal culture, for the establishment of a case-control resource population. A total of four SNPs viz. rs208222804, rs211654540, rs208814257, and rs210748127 in CD209 gene were genotyped by PCR-RFLP. All SNPs, except rs210748127, were polymorphic in our population. Genotypic-phenotypic associations were assessed by the PROCLOGISTIC procedure of SAS 9.3. The SNP rs208814257 yielded three genotypes viz. CC, CG, and GG, which were significantly (p < 0.05) different in case as compared to the control population. The odds of CC and CG in comparison to GG genotype were 1.21 and 0.40, respectively. The CG genotype was significantly higher in control population, indicating that this genotype may provide resistance against PTB in our resource population. Upon validation in an independent, larger test population and following biological characterization, SNP rs208814257 can be incorporated in marker panel for selection of animals with greater resistance to MAP infection.

Bayesian analysis reveals the influence of maternal effect on pre-weaning body weights in Landlly piglets.

The present study was undertaken to estimate the (co)variance components and genetic parameters of body weights recorded in Landlly piglets from birth to weaning at weekly intervals (w0 to w6). The data pertained to body weights of 2462 piglets, born to 91 sires and 159 dams across different generations during a 7-year period from 2014 to 2020. Five animal models (I-V), differentiated by inclusion or exclusion of maternal effects with or without covariance between maternal and direct genetic effects, were fitted on the data using the Bayesian algorithm. The analyses were implemented by Gibbs sampling in the BLUPF90 program and Markov chain Monte Carlo (MCMC) methodology was used to draw samples of posterior distribution pertaining to (co)variance components. Based on deviance information criteria (DIC), model V with inclusion of direct additive genetic, direct maternal genetic and permanent environmental effect of dam as random factors along with covariance between direct additive and maternal effects best fitted the data on pre-weaning traits (w0 to w5). Whereas, model I incorporating only the direct additive genetic effect best fitted the weaning weight (w6) data in Landlly piglets. The posterior mean estimates of direct heritability under the best models for W0 to W6 were 0.13, 0.19, 0.29, 0.13, 0.26, 0.32 and 0.46, respectively. Inclusion of the maternal component helped in better partitioning of variance for different body weights in Landlly piglets. The maternal heritability ranged from 0.06 to 0.14, while the litter heritability ranged from 0.11 to 0.15 for pre-weaning weights (W0 to W5) under the best-fit models. The influence of maternal environment was greater than maternal genetic effect from birth to 4th week of age. The results implied that variations in body weight of Landlly pigs were genetically controlled to moderate levels (especially w2 and w4) with contributions from direct additive and maternal genotype that can be exploited by designing efficient breeding programmes.

Exploration of allelic variants in short tandem repeats (STRs) flanking milk production QTLs and their association with milk production traits in Indian water buffaloes.

Short tandem repeats (STRs) are co-dominant, highly polymorphic marker loci, distributed throughout the genome and useful for identification and mapping of QTLs associated with variation in traits of economic significance. Allelic variants were identified in STRs located in close vicinity of cattle QTLs for milk production,, viz. BMS713, BM6404, BM4513, BM121, BM6105, TGLA245, BL1100, BMS1948, BMS711, BM1443, BM1706, BM6438, BM143, BM415, ETH131, ETH 2, and BM1329 in 109 water buffaloes of Murrah breed. All loci except TGLA245 exhibited polymorphism of varying degree. The observed number of alleles, effective number of alleles, PIC value, observed heterozygosity, and expected heterozygosity across all STR loci averaged 4.12 ± 0.22, 3.20 ± 0.22, 0.60 ± 0.04, 0.34 ± 0.05, and 0.66 ± 0.03, respectively. Goodness of fit (chi-square) and likelihood ratio (G square) test demonstrated that the population exhibited a deviation from HWE for all the loci. FIS was positive and ranged from 0.22 to 1.00. Least square analysis of variance exhibited significant effects of BM4513, ETH131, BM713, and BM6105 on first lactational total lactation milk yield, 305-day milk yield, lactation length, and dry period respectively. None of the STRs could exhibit significant effect on peak yield.

Comparative analysis of five different methods to design a breed-specific SNP panel for cattle.

Single nucleotide polymorphisms (SNPs) have now replaced microsatellite markers in several species for various genetic investigations like parentage assignment, genetic breed composition, assessment for individuality and, most popularly, as a useful tool in genomic selection. However, such a resource, which can offer to assist breed identification in a cost-effective manner is still not explored in cattle breeding programs. In our study, we have tried to describe methods for reducing the number of SNPs to develop a breed-specific panel. We have used SNP data from Dryad open public access repository. Starting from a global dataset of 178 animals belonging to 10 different breeds, we selected five panels each comprising of similar number of SNPs using different methods i.e., Delta, Pairwise Wright's FST, informativeness for assignment, frequent item feature selection (FIFS) and minor allele frequency-linkage disequilibrium (MAF-LD) based method. MAF-LD based method has been recently developed by us for construction of breed-specific SNP panels. The STRUCTURE software analysis of MAF-LD based method showed appropriate clustering in comparison to other panels. Later, the panel of 591 breed-specific SNPs was called to their respective breeds using Venny 2.1.0 and UGent web tools software. Breed-specific SNPs were later annotated by using various Bioinformatics softwares.

Review of Approaches for Increasing Ophthalmic Bioavailability for Eye Drop Formulations.

Ophthalmic diseases represent a significant problem as over 2 billion people worldwide suffer from vison impairment and blindness. Eye drops account for around 90% of ophthalmic medications but are limited in success due to poor patient compliance and low bioavailability. Low bioavailability can be attributed to short retention times in the eye caused by rapid tear turnover and the difficulty of drug diffusion through the multi-layered structure of the eye that includes lipid-rich endothelial and epithelial layers as well as the stroma which is high in water content. In addition, there are barriers such as tight junctional complexes in the corneal epithelium, lacrimal turnover, nasolacrimal drainage, blinking reflexes, efflux transporters, drug metabolism by ocular enzymes, and drug binding to or repulsion from conjunctival mucins, tear proteins, and melanin. In order to maximize transport through the cornea while minimizing drug loss through other pathways, researchers have developed numerous methods to improve eye drop formulations including the addition of viscosity enhancers, permeability enhancers, mucoadhesives, and vasoconstrictors, or using formulations that include puncta occlusion, nanocarriers, or prodrugs. This review explains the mechanism behind each of these methods, examines their history, analyzes previous and current research, evaluates future applications, and discusses the pros and cons of each technique.

Accurate assessment of body weights using morphometric measurements in Landlly pigs.

The aim of the study was to evaluate 16 novel morphometric traits of Landlly piglets at weaning (6 weeks) and post weaning (8 weeks) stage and to predict corresponding body weight from the measurements. A total of 279 Landlly piglets (n = 279, 75% Landrace + 25% Ghurrah crosses) were enrolled in this study. Body length, heart girth, paunch girth, height at wither, height at back, rump width, thigh circumference, neck circumference, and body depth had high correlation coefficients (0.8-0.97) with body weight at both the stages. Stepwise regression showed that body length and heart girth contributed most in prediction of both body weights while height at wither for body weight at 6 weeks and neck circumference for body weight at 8 weeks was the next highest contributing trait. Akaike's information criterion, Bayesian information criteria, adjusted R2, concordance correlation coefficient, bias correction factor, modeling efficiency, and coefficient of model determination were used to determine the most appropriate model for the prediction of body weight. Model containing body length and heart girth was fitted best to data for prediction of body weight at both weaning and post weaning stage with adjusted R2 values of 0.94 and 0.96, respectively. Hence, 2 different models were proposed for accurately predicting body weight in Landlly pigs at 6 and 8 weeks.

Population structure and admixture analysis in Frieswal crossbred cattle of India - a pilot study.

The present study was aimed to assess parameters related to genetic diversity, population structure and admixture in the Frieswal crossbred cattle of India. A total of three datasets were analyzed during this study. Dataset A (n = 80) consisted of data on two purebred populations, i.e., Shorthorn (n = 35) and Brahman (n = 25) and one crossbred strain Santa Gertrudis (n = 20). The dataset B (n = 71) consisted of data on three populations that included Holstein-Friesian (n = 30), Sahiwal (n = 27) and Frieswal (n = 14) cattle. The dataset C included data on all the six breeds under study. Dataset C was used to assess the indices of population structure and genetic diversity of different breeds prior to and after LD pruning. For Frieswal cattle strain, the proportion of polymorphic SNPs and MAF levels was 84.54% and 0.24, respectively. Frieswal strain maintained appreciable genetic diversity with observed heterozygosity measure of 0.414. PCA plots for three datasets depicted effective stratification of different breeds in the respective datasets. The genomic clustering levels of Sahiwal and Holstein-Friesian were found to be 98.45 and 99.89%, respectively, while the admixture of Frieswal was estimated to be about 61.5 and 38.5% from Holstein-Friesian and Sahiwal breeds, respectively.

Contact Lens Based Drug Delivery to the Posterior Segment Via Iontophoresis in Cadaver Rabbit Eyes.

PURPOSE: A drug loaded contact lens combined with electrodes positioned diametrically opposite and beyond the limbus can potentially deliver ionic drugs directly to the vitreous. METHODS: Commercial lenses are loaded with nile blue or fluorescein as the drug analogs and placed on cadaver rabbit eyes. Electrodes (19.6 mm2) are placed atop at opposite sides of the sclera to apply a constant current (0.125-0.250 mA) for 1-2 h. COMSOL simulations are conducted to determine the field distribution and the potential drop across various tissue layers and equivalent circuit model is developed to calculate the electrophoretic velocity and estimate the drug flux. RESULTS: The device delivered both hydrophobic and hydrophilic dyes to the tissue. The amount of fluorescein delivered to the vitreous directly correlated with the applied current and time duration. The electrophoretic mobility from the experimental data agreed with the model estimates. Confocal microscopy showed that nile blue penetrated through the conjunctiva-sclera barrier to reach the retina showing that the electric field can transport molecules through the ocular tissue and into the vitreous. The ex vivo model neglects transport into flowing capillaries in the choroid. However, the time scale for electrophoretic transport across the choroid was found to be 550-1300 fold shorter than that for uptake into the choroidal capillaries. CONCLUSION: Incorporation of an electric field with multiple electrodes on a single lens can effectively deliver ionic drugs to the posterior region at levels comparable to current methods with the benefits of being safer and less invasive.

Cloning and characterization of Bubaline mammary miRNAs: An in silico approach.

MicroRNAs (miRNAs) are small non-coding RNAs ~ 19-25 nucleotides long that are involved in the regulation of gene expression. They negatively regulate the gene expression via inhibition or complete degradation of mRNAs by binding the complementary target sequences in 3' untranslated region. The present investigation was aimed at profiling of miRNAs expressed in the Bubaline mammary tissue at dry stage of lactation cycle. Small RNAs were isolated from freshly collected mammary tissues and T4 RNA ligase was used to ligate the enriched miRNAs with 3' and 5' linker sequences in two separate reactions. cDNA copies were synthesized from linkered small RNAs followed by the PCR amplification. The PCR products were resolved on 15% non-denaturing polyacrylamide gel electrophoresis by gelstar staining. The PCR products were cloned using pGEM®-T easy vector system and the desired clones (with linkered small RNA sequences) were confirmed using restriction digestion of plasmids with EcoRI. Out of 15 Bubaline small RNA sequences, eight sequences (Seq. ID I-VIII) matched the size range of miRNA molecules i.e., 18-26 nucleotides. The Bubaline small RNA sequences II and III showed partial alignment with various mammalian and non-mammalian miRNAs. The small RNA sequences obtained in the present study did not show any perfect match with already reported mRNA, rRNA or tRNA sequences in different databases. Hence, only the Bubaline small RNA sequences that showed partial homology with miRNAs were considered as putative Bubaline miRNAs. The present study established the basic repertoire of miRNAs expressed at dry stage of lactation in Bubaline mammary gland.