Characterization of 24bp Insertion Polymorphism of Prolactin Gene and its Association with Quantitative Traits in Tellicherry Native Chicken Breed

Background: Over many centuries, the conventional methods of poultry breeding is based on the phenotypic selection of the birds with superior trait in a particular population. Prolactin is a physiological candidate gene that has significant effects on production traits in poultry. The objective of this study was to investigate 24bp insertion (I) polymorphism of prolactin gene and its association with quantitative traits in Tellicherry native chicken. Methods: A total of 200 blood samples were collected from the randomly selected birds of All India Co-ordinated Research Project (AICRP) on Poultry Breeding, Mannuthy. The isolated DNA samples were subjected to polymerase chain reaction (PCR) by using specific set of primers to amplify the 24bp insertion polymorphism. PCR amplicons were sequenced to study the polymorphism at nucleotide level. Result: PCR amplification revealed the presence of a specific DNA fragment with 130/154bp contained 24bp insertion polymorphism. Based on the polymorphic patterns birds were designated with three genotypes as II, ID and DD. The frequency of I (0.6975) allele was found higher than D (0.3025) allele. Our experimental results revealed that 24bp insertion polymorphism of prolactin did not have a significant association with production traits viz, age at sexual maturity, egg weight and egg number in Tellicherry native chicken.

Assessment of genetic diversity and variety identification based on developed retrotransposon-based insertion polymorphism (RBIP) markers in sweet potato (Ipomoea batatas (L.) Lam.)

Sweet potato, a dicotyledonous and perennial plant, is the third tuber/root crop species behind potato and cassava in terms of production. Long terminal repeat (LTR) retrotransposons are highly abundant in sweet potato, contributing to genetic diversity. Retrotransposon-based insertion polymorphism (RBIP) is a high-throughput marker system to study the genetic diversity of plant species. To date, there have been no transposon marker-based genetic diversity analyses of sweet potato. Here, we reported a structure-based analysis of the sweet potato genome, a total of 21555 LTR retrotransposons, which belonged to the main LTR-retrotransposon subfamilies Ty3-gypsy and Ty1-copia were identified. After searching and selecting using Hidden Markov Models (HMMs), 1616 LTR retrotransposon sequences containing at least two models were screened. A total of 48 RBIP primers were synthesized based on the high copy numbers of conserved LTR sequences. Fifty-six amplicons with an average polymorphism of 91.07% were generated in 105 sweet potato germplasm resources based on RBIP markers. A Unweighted Pair Group Method with Arithmatic Mean (UPGMA) dendrogram, a model-based genetic structure and principal component analysis divided the sweet potato germplasms into 3 groups containing 8, 53, and 44 germplasms. All the three analyses produced significant groupwise consensus. However, almost all the germplasms contained only one primary locus. The analysis of molecular variance (AMOVA) among the groups indicated higher intergroup genetic variation (53%) than intrapopulation genetic variation. In addition, long-term self-retention may cause some germplasm resources to exhibit variable segregation. These results suggest that these sweet potato germplasms are not well evolutionarily diversified, although geographic speciation could have occurred at a limited level. This study highlights the utility of RBIP markers for determining the intraspecies variability of sweet potato and have the potential to be used as core primer pairs for variety identification, genetic diversity assessment and linkage map construction. The results could provide a good theoretical reference and guidance for germplasm research and breeding.

The pan HDAC inhibitor Givinostat improves muscle function and histological parameters in two Duchenne muscular dystrophy murine models expressing different haplotypes of the LTBP4 gene

Background In the search of genetic determinants of Duchenne muscular dystrophy (DMD) severity, LTBP4, a member of the latent TGF-β binding protein family, emerged as an important predictor of functional outcome trajectories in mice and humans. Nonsynonymous single-nucleotide polymorphisms in LTBP4 gene associate with prolonged ambulation in DMD patients, whereas an in-frame insertion polymorphism in the mouse LTBP4 locus modulates disease severity in mice by altering proteolytic stability of the Ltbp4 protein and release of transforming growth factor-β (TGF-β). Givinostat, a pan-histone deacetylase inhibitor currently in phase III clinical trials for DMD treatment, significantly reduces fibrosis in muscle tissue and promotes the increase of the cross-sectional area (CSA) of muscles in mdx mice. In this study, we investigated the activity of Givinostat in mdx and in D2.B10 mice, two mouse models expressing different Ltbp4 variants and developing mild or more severe disease as a function of Ltbp4 polymorphism. Methods Givinostat and steroids were administrated for 15 weeks in both DMD murine models and their efficacy was evaluated by grip strength and run to exhaustion functional tests. Histological examinations of skeletal muscles were also performed to assess the percentage of fibrotic area and CSA increase. Results Givinostat treatment increased maximal normalized strength to levels that were comparable to those of healthy mice in both DMD models. The effect of Givinostat in both grip strength and exhaustion tests was dose-dependent in both strains, and in D2.B10 mice, Givinostat outperformed steroids at its highest dose. The in vivo treatment with Givinostat was effective in improving muscle morphology in both mdx and D2.B10 mice by reducing fibrosis. Conclusion Our study provides evidence that Givinostat has a significant effect in ameliorating both muscle function and histological parameters in mdx and D2.B10 murine models suggesting a potential benefit also for patients with a poor prognosis LTBP4 genotype.

Detection of Deletion/Insertion Polymorphism Profiles from Single Human Hair Shafts

Background: Hair is a frequently encountered biological evidence in personal identification. The amount of nuclear DNA that can be extracted from a single strand of rootless hair is most limited, making the detection of short tandem repeat (STR) polymorphisms difficult. To overcome these limitations, deletion/insertion polymorphisms (DIP) as a new type of genetic marker have shown their benefits in detecting low-copy-number DNA. The Investigator DIPplex kit contains 30 biallelic autosomal DIP and amelogenin. The analysis of DIPs combines the advantages of both STR and single nucleotide polymorphism analyses. Thus, this study aimed to detect the DIP distribution of individual hair shafts from individuals.Methods and Results: DNA was extracted from the shaft of fresh, aged, and shed hair. After DNA was evaluated, the DIP profiles were detected by capillary electrophoresis. The results indicated that the amount of DNA extracted from hair roots was much higher than that from the hair shafts in the same individual for all samples. The degradation index values of DNA from the aged hair shafts were highest. It is classified to be “mildly degraded.” Compared with their hair roots, the full DIP profiles were detected for fresh hair, 70% for aged hair, and 92% for shed hair. Contrarily, except for fresh hair shafts, only three STR loci of the aged and shed strands of hair could be amplified using AmpFlSTR MiniFiler PCR Amplification Kit.Conclusions: These results indicate that the detection of DIP profile is an effective tool for personal identification from hair shafts, including aged hair.

Assessment of Genetic Diversity and Variety Identification Based on Developed Retrotransposon-Based Insertion Polymorphism (RBIP) Markers in Sweet Potato (Ipomoea Batatas (L.) Lam)

Sweet potato, a dicotyledonous and perennial plant, is the third most crucial tuber/root crop species behind potato and cassava in terms of production. Long terminal repeat (LTR) retrotransposons are highly abundant in sweet potato, contributing to genetic diversity. These LTR retrotransposons play a significant role in sweet potato genotypes. Retrotransposon-based insertion polymorphism (RBIP) is a high-throughput marker system to study the genetic diversity of plant species. To date, there have been no transposon marker-based genetic diversity analyses of sweet potato. We report a structure-based analysis of the sweet potato genome for the main LTR-retrotransposon subfamilies, Ty3-gypsy and Ty1-copia, which revealed a total of 21555 LTR retrotransposons. By searching using hidden Markov models (HMMs), 1616 LTR retrotransposon sequences containing at least two models were found. A total of 48 RBIP primers were synthesized based on the high copy numbers of conserved LTR sequences. RBIP markers of the genetic diversity and population structure of 105 sweet potato germplasm resources revealed 56 amplicons with an average polymorphism of 91.07%. Sweet potato accessions were collected from 6 provinces of China, Japan and America. A UPGMA dendrogram, a model-based genetic structure and principal component analysis (PCA) divided the sweet potato germplasms into 3 groups containing 8, 53, and 44 germplasms. All three analyses produced significant groupwise consensus. However, almost all the germplasms contained only one primary locus. The analysis of molecular variance (AMOVA) among the groups indicated higher intergroup genetic variation (53%) than intrapopulation genetic variation. In addition, long-term self-retention may cause some germplasm resources to exhibit variable segregation. These results suggest that these sweet potato germplasms are not well evolutionarily diversified, although geographic speciation could have occurred at a limited level. This study highlights the utility of RBIP markers for determining the intraspecies variability of sweet potato.

A SINE-VNTR-Alu in the LRIG2 Promoter Is Associated with Gene Expression at the Locus

The hominid SINE-VNTR-Alu (SVA) retrotransposons represent a repertoire of genomic variation which could have significant effects on genome function. A human-specific SVA in the promoter region of the gene leucine-rich repeats and immunoglobulin-like domains 2 (LRIG2), which we termed SVA_LRIG2, is a common retrotransposon insertion polymorphism (RIP), defined as an element which is polymorphic for its presence or absence in the genome. We hypothesised that this RIP might be associated with differential levels of expression of LRIG2. The RIP genotype of SVA_LRIG2 was determined in a subset of frontal cortex DNA samples from the North American Brain Expression Consortium (NABEC) cohort and was imputed for a larger set of that cohort. Utilising available frontal cortex total RNA-seq and CpG methylation data for this cohort, we observed that increased allele dosage of SVA_LRIG2 was non-significantly associated with a decrease in transcription from the region and significantly associated with increased methylation of the CpG probe nearest to SVA_LRIG2, i.e., SVA_LRIG2 is a significant methylation quantitative trait loci (mQTL) at the LRIG2 locus. These data are consistent with SVA_LRIG2 being a transcriptional regulator, which in part may involve epigenetic modulation.

Phylogenetic relationships among accessions in Citrus and related genera based on the insertion polymorphism of the CIRE1 retrotransposon

A total of 145 accessions of the genus Citrus and related genera, maintained in the Conservation Garden for Citrus Germplasm at the Experimental Farm of Kindai University, Yuasa, Wakayama, Japan, were examined for their phylogenetic relationships. The present classification was conducted using an inter-retrotransposon amplified polymorphism (IRAP) method based on the insertion polymorphism of a retrotransposon, CIRE1, identified in C. sinensis. The objective of this study was to evaluate the applicability of the IRAP method for citrus classification. The constructed dendrogram showed that the 145 accessions and two outgroup species were successfully classified into five major clades. A large number of C. sinensis accessions were divided into three traditional groups, navel orange, sweet orange, and blood orange, almost corresponding to the sub-clades in the dendrogram. Several other accessions belonging to the same species, and also many hybrid cultivars from crossbreeding, were localized into the respective sub-clades or near positions in the dendrogram. Several unclassified accessions could also be located in the dendrogram, suggesting novel relationships with other accessions. It was concluded that the IRAP method based on CIRE1 insertion polymorphism was suitable for the classification of citrus from a molecular point of view.