Single Nucleotide Polymorphism Mutation Research Articles

Molecular characterization of varicella zoster virus isolated from clinical samples in India.

Background & objectives:Varicella zoster virus (VZV) strains are classified into six different clades based on the sequencing of its genome. Clades 4 and 5 are reported from India based on the single-nucleotide polymorphism (SNP). Till now, multiple clade circulations using partial sequences have been reported from India due to the lack of availability of the full VZV genome sequence. This study conducted a genome sequencing of VZV in India to identify circulating clade.Methods:Four clinical samples obtained from symptomatic patients tested positive for VZV by real-time PCR were used. These four samples were preferred to retrieve the genomic VZV sequence using the next-generation sequencing method. A reference-based assembly method was used to retrieve the genome of VZV, which was further analyzed.Results:At the least, 98 per cent of the whole-genome sequences were recovered from the four samples. The VZV sequences obtained in this study formed a separate monophyletic branch with clade 5, indicating it to be evolved from a distinct ancestor. The nucleotide-based analysis revealed 13 different SNP mutations and one multiple nucleotide variation in the VZV sequences when compared to one of the clade 5 genomes having accession number: DQ457052.1.Interpretation & conclusions:The present study described approximately 98 per cent of the genome sequence of VZV from India. The availability of these genomic sequences will lead to enrichment in the clinical genomic data set from India. The available data would help in the development of diagnostic methods along with evolutionary analysis. We hypothesize the existence of a new sub-clade that belongs to clade 5 and propose further experiments to confirm these results.

Population genomic transformations induced by isolation of wild bird avian influenza viruses (Orthomyxoviridae) in embryonated chicken eggs.

Isolation and cultivation of wild-type viruses in model organism cells or tissues is standard practice in virology. Oftentimes, the virus host species is distantly related to the species from which the culture system was developed. Thus, virus culture in these tissues and cells basically constitutes a host jump, which can lead to genomic changes through genetic drift and/or adaptation to the culture system. We directly sequenced 70 avian influenza virus (Orthomyxoviridae) genomes from oropharyngeal/cloacal swabs collected from wild bird species and paired virus isolates propagated from the same samples following isolation in specific-pathogen-free embryonated chicken eggs. The data were analyzed using population genetic approaches including evaluation of single nucleotide polymorphism (SNP) frequencies and divergence with pooled-sequencing analyses, consensus sequence placement in neighbor-joining trees, and haplotype reconstruction and networks. We found that propagation of virus in eggs leads to skewed SNP mutation spectra with some SNPs going to fixation. Both synonymous and nonsynonmous SNP frequencies shifted. We found multiple consensus sequences that differed between the swabs and the isolates, with some sequences from the same sample falling into divergent genetic clusters. Twenty of 23 coinfections detected had different dominant subtypes following virus isolation, thus sequences from both the swab and isolate were needed to obtain full subtype data. Haplotype networks revealed haplotype frequency shifts and the appearance or loss of low-frequency haplotypes following isolation. The results from this study revealed that isolation of wild bird avian influenza viruses in chicken eggs leads to skewed populations that are different than the input populations. Consensus sequence changes from virus isolation can lead to flawed phylogenetic inferences, and subtype detection is biased. These results suggest that for genomic studies of wild bird influenza viruses the biological field should move away from chicken egg isolation towards directly sequencing the virus from host samples.

Screening and identification of biomarkers associated with the diagnosis and prognosis of lung adenocarcinoma.

BackgroundIn this study, we aimed to identify the pathogenesis and prognostic biomarkers of lung adenocarcinoma (LUAD).MethodsDifferentially expressed mRNAs (DEmRNAs) and single nucleotide polymorphism (SNP) mutant genes were screened. In addition, enrichment and protein‐protein interaction (PPI) network analyses of the SNP‐mutated genes were performed. Thereafter, the correlation between gene mutation and expression was analyzed. Finally, the mutated genes associated with LUAD prognosis were validated on the basis of The Cancer Genome Atlas (TCGA) database.ResultsA total of 2502 DEmRNAs were initially screened in this study. We identified 756 SNP‐mutated genes from more than 30 cases. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that the mutated genes involved in LUAD were mainly associated with the ECM‐receptor interaction, focal adhesion, and calcium signaling pathways. Tumor protein p53 (TP53) and neurexin 1 (NRXN1) with the higher degree were chosen as the hub genes in the PPI network. In addition, the correlation analysis revealed six genes, including assembly factor for spindle microtubules (ASPM), centromere protein F (CENPF), contactin 3 (CNTN3), catenin delta 2 (CTNND2), PKHD1 like 1 (PKHD1L1), and semaphorin 6D (SEMA6D), and three SNP mutations at ASPM rs368020495, CENPF rs762653487, and PKHD1L1 rs768349010 sites that were found to be associated with LUAD prognosis. Further validation showed that among the aforementioned six mutated genes, CENPF was upregulated and SEMA6D was downregulated.Conclusion CENPF, SEMA6D, TP53, and NRXN1 were found to be closely associated with the development of LUAD.

The complete chloroplast genome of Isochrysis galbana and comparison with related haptophyte species

The haptophyte Isochrysis galbana is an algal species of commercial interest due to its ability to accumulate high levels of fucoxanthin. In this study, the complete chloroplast (cp) genome of I. galbana was sequenced using a combination of Illumina and third-generation sequencing platforms. This circular cp genome was 105,872 bp in length, consisting of a large single-copy (LSC) region of 54,838 bp and a small single-copy (SSC) region of 41,308 bp separated by two direct rDNA repeats of 4862 bp and 4864 bp. The cp genome contained 141 unique genes, including 112 protein-coding genes with no introns, 26 transfer-RNA (tRNA) genes and three ribosomal-RNA (rRNA) genes. Like canonical ribosomal operon repeat structure, two 16S-trnI-trnA-23S-5S ribosomal RNA operons were identified. A total of 32 pairs of small repeats and 45 perfect microsatellites were detected. All haptophyte chloroplasts sequenced to date apart from E. huxleyi have either a complete pair of canonical repeats with direct orientation or noncanonical inverted repeats that lost part of tRNAs, or they have lost the inverted repeat arrangement altogether. Additionally, imperfect ribosomal operon repeats were observed in most haptophytes. The non-identical 4.9 kb ribosomal direct-repeats in I. galbana cp genome were distinguished by three 1–2 bp-sized inserts/deletions (InDels) and 32 single nucleotide polymorphisms (SNPs). Pairwise chloroplast genome comparisons showed a high degree of conservation in genome structure, gene order and gene content between I. galbana and T. lutea, with an overall SNP mutation rate of 4.3 × 10−3 per site. Only 458 SNPs, 18 small InDels and one large insertion were detected between these two cp genomes. The transition-to-transversion (Ts/Tv) ratio was 1.79 and the predominant mutations were A/T to G/C transitions. Phylogenetic analysis of currently available haptophyte chloroplasts also reflected that I. galbana was most closely related to T. lutea. The Ka/Ks ratios of shared genes in Isochrysidales chloroplast genomes were less than 1, suggesting that those genes are under purifying selection. The phylogenetic tree yielded by plastome single-copy orthologous genes gives new evidence on the relationships among CASH lineages. This first I. galbana chloroplast genome will provide insight into the chloroplast architecture, function and evolution of this species and other haptophytes.

Single Nucleotide Polymorphisms in the Promoter of CYP19 Gene in Cattle Bred in Iraq

The present study was undertaken to characterize the genetic diversity of the aromatase cytochrome P450 (CYP19) gene in 34 cows (15 local, 14 Holstein, and 5 Crosses) in Iraq. The objectives of the present study are to detect SNPs (mutations) in promoter p1.1 of the CYP19 gene in cattle bred in Iraq using sequencing techniques. We identified five single-nucleotide polymorphisms (SNP) loci of the CYP19 gene that were detected, namely G933T, G994C, A1044G, A1062T, and C1468A. The results showed the presence of 3, 4, and 2 polymorphic sites leading to the construction of 4, 5, and 3 different haplotypes for Holstein, local, and crosses respectively. Haplotype diversity were 0.791, 0.752, and 0.700 respectively. While nucleotide diversity was 0.0017, 0.0022, and 0.0013 respectively. Besides, we carried out a phylogenetic analysis of these sequences to address the evolutionary relationship between the animal species. These fragments were assigned in the GenBank database under the accession numbers: LC490756, LC490757, LC491437, LC491438, LC491439, LC491588, and LC491589.

Genomic variation between PRSV resistant transgenic SunUp and its progenitor cultivar Sunset

BackgroundThe safety of genetically transformed plants remains a subject of scrutiny. Genomic variants in PRSV resistant transgenic papaya will provide evidence to rationally address such concerns.ResultsIn this study, a total of more than 74 million Illumina reads for progenitor ‘Sunset’ were mapped onto transgenic papaya ‘SunUp’ reference genome. 310,364 single nucleotide polymorphisms (SNPs) and 34,071 small Inserts/deletions (InDels) were detected between ‘Sunset’ and ‘SunUp’. Those variations have an uneven distribution across nine chromosomes in papaya. Only 0.27% of mutations were predicted to be high-impact mutations. ATP-related categories were highly enriched among these high-impact genes. The SNP mutation rate was about 8.4 × 10− 4 per site, comparable with the rate induced by spontaneous mutation over numerous generations. The transition-to-transversion ratio was 1.439 and the predominant mutations were C/G to T/A transitions. A total of 3430 nuclear plastid DNA (NUPT) and 2764 nuclear mitochondrial DNA (NUMT) junction sites have been found in ‘SunUp’, which is proportionally higher than the predicted total NUPT and NUMT junction sites in ‘Sunset’ (3346 and 2745, respectively). Among all nuclear organelle DNA (norgDNA) junction sites, 96% of junction sites were shared by ‘SunUp’ and ‘Sunset’. The average identity between ‘SunUp’ specific norgDNA and corresponding organelle genomes was higher than that of norgDNA shared by ‘SunUp’ and ‘Sunset’. Six ‘SunUp’ organelle-like borders of transgenic insertions were nearly identical to corresponding sequences in organelle genomes (98.18 ~ 100%). None of the paired-end spans of mapped ‘Sunset’ reads were elongated by any ‘SunUp’ transformation plasmid derived inserts. Significant amounts of DNA were transferred from organelles to the nuclear genome during bombardment, including the six flanking sequences of the three transgenic insertions.ConclusionsComparative whole-genome analyses between ‘SunUp’ and ‘Sunset’ provide a reliable estimate of genome-wide variations and evidence of organelle-to-nucleus transfer of DNA associated with biolistic transformation.

Polymorphisms in the ASMT and ADAMTS1 gene may increase litter size in goats

Prolificacy of most local goat breeds in China is low. Jining Grey goat is one of the most prolific goat breeds in China, it is an important goat breed for the rural economy. ASMT (acetylserotonin O‐methyltransferase) and ADAMTS1 (ADAM metallopeptidase with thrombospondin type 1 motif) are essential for animal reproduction. Single nucleotide polymorphisms (SNPs) of ASMT and ADAMTS1 genes in the highly prolific breed (Jining Grey goats), medium prolific breed (Boer goats and Guizhou White goats) and low prolific breeds (Angora goats, Liaoning Cashmere goats and Inner Mongolia Cashmere goats) were detected by polymerase chain reaction‐restriction fragment length polymorphism and sequencing. Two SNPs (g.158122T>C, g.158700G>A) of ASMT gene and two SNPs (g.7979798A>G, g.7979477C>T) of ADAMTS1 gene were identified. For g.158122T>C of ASMT gene, further analysis revealed that genotype TC or CC had 0.66 (p < 0.05) or 0.75 (p < 0.05) kids more than those with genotype TT in Jining Grey goats. No significant difference (p > 0.05) was found in litter size between TC and CC genotypes. The SNP (g.158122T>C) caused a p.Tyr298His change and this SNP mutation resulted in changes in protein binding sites and macromolecule‐binding sites. The improvement in reproductive performance may be due to changes in the structure of ASMT protein. For g.7979477C>T of ADAMTS1 gene, Jining Grey does with genotype CT or TT had 0.82 (p < 0.05) or 0.86 (p < 0.05) more kids than those with genotype CC. No significant difference (p > 0.05) was found in litter size between CT or TT genotypes. These results preliminarily indicated that C allele (g.158122T>C) of ASMT gene and T allele (g.7979477C>T) of ADAMTS1 gene are potential molecular markers which could improve litter size of Jining Grey goats and be used in goat breeding.

Mitochondrial genome variation between different accessions of Matricaria chamomilla L. (Asteraceae) based on SNP mutation analysis

Matricaria chamomilla L. (chamomile, Asteraceae) (GRIN, The Plant List 2013) has a long history of usage in traditional herbal medicine and is still today amongst the most important medicinal plants. Despite this importance, genetic diversity of cultivated and wild germplasm of M. chamomilla was rarely investigated so far. The objective of this study was to estimate the mitochondrial (mt) diversity of various cultivated M. chamomilla genotypes by determining point mutations in the mt genome. 89 SNPs (single nucleotide polymorphisms) were identified in the next generation sequencing data of 33 genotypes from 11 di- and tetraploid chamomile accessions representing a sequence diversity of 0.32 SNPs/kb. Based on the SNP analysis 19 mitochondrial haplotypes (mitotypes) could be specified with genetic distances ranging between 0.011 and 0.851. The examined mt variability within the accessions was higher than expected; only one monomorphic accession (variety ‘Camoflora’) was identified. Diploid accessions exhibited with 1.9 mitotypes per accession a higher variability than tetraploid accessions with a ratio of 1.3. Although some of the mitotypes were distributed over different accessions, identical mitotypes within di- and tetraploid accessions could not be determined. Furthermore, the mitotypes did not correspond to the geographical origin of the accessions. Although not the whole mt genome could be assembled in this study, the substitutions identified represent a valuable tool for further investigations of maternal phylogenetic relationships within M. chamomilla.

Mapping a double flower phenotype-associated gene DcAP2L in Dianthus chinensis.

The double flower is a highly important breeding trait that affects the ornamental value in many flowering plants. To get a better understanding of the genetic mechanism of double flower formation in Dianthus chinensis, we have constructed a high-density genetic map using 140 F2 progenies derived from a cross between a single flower genotype and a double flower genotype. The linkage map was constructed using double-digest restriction site-associated DNA sequencing (ddRAD-seq) with 2353 single nucleotide polymorphisms (SNPs). Quantitative trait locus (QTL) mapping analysis was conducted for 12 horticultural traits, and major QTLs were identified for nine of the 12 traits. Among them, two major QTLs accounted for 20.7% and 78.1% of the total petal number variation, respectively. Bulked segregant RNA-seq (BSR-seq) was performed to search accurately for candidate genes associated with the double flower trait. Integrative analysis of QTL mapping and BSR-seq analysis using the reference genome of Dianthus caryophyllus suggested that an SNP mutation in the miR172 cleavage site of the A-class flower organ identity gene APETALA2 (DcAP2L) is responsible for double flower formation in Dianthus through regulating the expression of DcAG genes.

Mitochondrial D-loop informative SNPs in identification of dog’s breed

The aim of the study was to determine differences at the molecular level between different breeds of dogs on the basis of a comparative analysis of the mtDNA D-loop sequence. The material used in the study consisted of blood collected from 40 Doberman Pinscher, Dachshund, German Shepherd, and Crossbreed dogs. The investigations involved DNA extraction, amplification and sequencing of the mtDNA D-loop, as well as bioinformatic analyses. Fifteen SNP (single nucleotide polymorphism) mutations were identified. This study proposes a new solution for dog breed identification, i.e. the use of SNPs specific to each breed. The analysis revealed the presence of already known SNPs in various configurations depending on the breed. Breed-specific SNP patterns were identified in 94% of the Doberman dogs, and two SNP patterns were found in 73% of the German Shepherd dogs. This subject requires further research. However, given the limited amount of data on the use of mtDNA in determination of breed affiliation, the present results seem to be promising.

Two types of mutations in the HEUKCHEEM gene functioning in cucumber spine color development can be used as signatures for cucumber domestication.

The HEUKCHEEM gene plays an important role in spine color formation. A white spine occurs due to two mutations in HEUKCHEEM and is closely related to the regional distribution of these mutants. Mapping analysis revealed that the HEUKCHEEM gene is co-segregated with the B locus in the regulation of black spine color development in cucumber fruit. HEUKCHEEM induced the expression of the genes involved in the anthocyanin biosynthetic pathway, leading to the accumulation of anthocyanins in black spines. The transiently over-expressed HEUKCHEEM in cucumber and tobacco plants enhanced the expression of anthocyanin biosynthesis-related genes, leading to anthocyanin accumulation. However, two mutations-insertion of the 6994bp mutator-like transposable element (MULE) sequence into the second intron and one single-nucleotide polymorphism (SNP) of C to T in the second exon of HEUKCHEEM-were identified in white spines, leading to no accumulation of anthocyanin biosynthesis-related gene transcripts and anthocyanins. Furthermore, association analysis using 104 cucumber accessions with different geographical origins revealed that the types of mutations in HEUKCHEEM are strongly linked to geographical origins. The MULE insertion is found extensively in cucumbers with white spines in East Asia and Australia. However, cucumbers with white spines in other areas could be significantly influenced by a single SNP mutation. Our results provide fundamental information on spine color development in cucumber fruits and spine color-based cucumber breeding programs.

Highly sensitive and selective detection of single-nucleotide polymorphisms using gold nanoparticle MutS enzymes and a micro cantilever resonator

Mutant DNAs are important markers useful for the diagnosis of human disease. Single-nucleotide polymorphisms (SNPs) represent the most common types of DNA mutations. As there is only a one base pair change in a single nucleotide between the SNP and the wild-type DNA, it is difficult to distinguish the SNPs. In this report, a highly sensitive and selective detection and discrimination of SNPs is performed using MutS, gold nanoparticles (AuNP) and a resonator. A single mismatched base exists between the SNP mutation and the probe DNA on the resonator, and MutS binds to the DNA at the location of the mismatch. As MutS is attached to AuNP (MutS-AuNP), both MutS and AuNP are adsorbed onto the resonator. The detection is based on the resonance frequency shift of the resonator following the adsorption of MutS-AuNP on the resonator. Highly sensitive detection of DNA mutations was achieved using AuNPs that act as mass amplifiers, and the obtained limit of detected was 100 fM. Additionally, our proposed method detected mutations in the presence of as little as 0.1% wild-type, and discrimination of specific mutations was also achieved. The results obtained from our proposed method suggest its potential for diagnosing cancer patients.

Understanding the pathophysiology behind chairside diagnostics and genetic testing for IL-1 and IL-6.

In order for chairside diagnostic testing to make an impact on dental therapy, practitioners require a better understanding of genetic mutations contributing to the pathophysiology of periodontal disease. Commensal and pathogenic bacterial colonization in oral cavity tissues produces a cascade of proinflammatory signaling pathways ultimately detrimental to host tissues. Resolving inflammation is a multifactorial process involving the downregulation of proinflammatory cytokines while allowing commensal bacterial levels to return to normal. Because of the complicated nature of commensal bacteria and oral health homeostasis, the emphasis of dental therapy should place renewed focus on limiting destructive inflammation rather than solely eliminating bacteria. Salivary diagnostics are an easy, non-invasive way to assess inflammatory markers. Inflammatory cytokine levels can help determine the subclinical health of a patient, showing the transition from health to gingivitis, or periodontitis, prior to clinical presentation. Single nucleotide polymorphism mutations can aid in determining increased risk of developing periodontitis. Taken together, and alongside regular clinical evaluations, chairside diagnostics help individualize treatment plans to slow, or halt, the progression of disease-before tissue destruction can take place. While more studies are needed analyzing specific mutations across periodontal categories, chairside diagnostics present an exciting adjunct to improve patient care.

Clinical significance of bone morphogenetic protein in osteosarcoma: A systematic review

Background: The literature suggests that endogenous bone morphogenetic proteins (BMP) have both beneficial and deleterious effects on osteosarcoma. Therapeutic use of exogenous BMP after osteosarcoma resection to enhance skeletal reconstruction is controversial. Methods: We conducted a systematic review to determine the association of endogenous BMP expression and overall survival for osteosarcoma patients to determine the association of BMP expression and metastasis, and to evaluate the extent of clinical evidence regarding use of recombinant human BMP (rhBMP) in patients with osteosarcoma. Results: Eight studies were included; all except one reported on endogenous BMPs or BMP receptor. The four clinical studies on an association between BMPs and prognosis had mixed results; one associated BMP-7 expression with improved overall survival (OS) in osteosarcoma, while another found no correlation between BMP-6, 7, or 8 with OS. In contrast, one found a negative correlation between BMP expression and prognosis. The final study of these four investigated BMP-2 single nucleotide polymorphisms (SNP) in osteosarcoma and reported an association of a specific SNP mutation with improved prognosis. All three studies addressing metastasis reported increased risk for metastasis of osteosarcoma with either BMP-2 receptor or BMP-2 or 4 expression. Only one study evaluated application of therapeutic rhBMP-2 after osteosarcoma resection, finding no effect on clinical outcomes and no detrimental effects such as tumor recurrence. Conclusions: Endogenous BMP or BMP receptor expression may be associated with osteosarcoma metastasis but are of uncertain prognostic significance. Any harmful or beneficial effects of exogenous therapeutic BMP application in this setting are unproven.

Candidate mutations used to aid the prediction of genetic merit for female reproductive traits in tropical beef cattle

In this study, we aimed to provide a wet laboratory validation for a set of single nucleotide polymorphisms (SNP), which had been identified as candidate functional variants in silico. Genotyping for candidate SNP was performed in Brahman and Tropical Composite cattle. After quality control, 29 SNP were first investigated individually for their association with female reproductive traits and then used as a panel for genomic predictions. The reproductive traits studied were age at first corpus luteum (AGECL; days), post-partum anoestrus interval (PPAI; days), and a binary trait that described if the cow had ovulated before weaning the first calf or not (PW, 0-1). Single nucleotide polymorphisms in six genes (FOXA2, TRAF4, IRF2, IRF1, BPTF, and CPEB1) were found to be significantly associated with reproduction traits . The genomic prediction method used was BayesR, to accommodate the 29 new SNP and compare their performance with predictions based on 50K genotypes (Illumina SNP chip). When new SNP and PLAG1 mutation rs109231213 were included in the genomic predictions for female reproductive traits their accuracies improved. The best predictions were obtained by combining the new SNP and the 50K SNP using BayesR analysis, with a 4% improvement in accuracy. The proportion of the genetic variance explained by the new SNP together was 0.07 for AGECL, 0.03 for PPAI, and 0.02 for PW. It would be favourable to include these new SNP in future versions of bovine SNP chips to target selection for female reproductive traits. These new SNP are likely to improve genomic predictions for female reproductive traits in tropical beef cattle breeds, with varying degrees of Bos indicus content.

Identification of polymorphism in the bovine interleukin-17A gene and its association with mastitis in Polish Holstein–Friesian cattle

The main aim of this study was to identify the association between the single nucleotide polymorphism (SNP) in the bovine interleukin-17A (IL-17A) gene and mastitis resistance. Additionally, the connection between the identified polymorphism and the somatic cell score (SCS) was tested. Two hundred and six Polish Holstein?Friesian cows were genotyped using the PCR-RFLP method. One SNP mutation (c.126G>A) was identified within the coding sequence of the analyzed gene. Results of logistic regression analysis displayed no significant association between the IL-17A genotype and clinical mastitis occurrence (Wald chi-square test = 0.74; P = 0.69). The GLM analysis demonstrated that genotype also did not affect the SCS (P = 0.77), while the effects of age at first calving, lactation number, lactation stage, mean milk yield per milking day, and milk yield on the day of milk recording proved to be highly significant (P < 0.01). The present study was one of the first to show an association between polymorphism within the IL-17A gene and selected udder health traits in cattle. An influence of the c.126G>A polymorphism on clinical mastitis and the SCS in the studied set of Polish Holstein?Friesian cows was not found. This suggests that the c.126G>A marker is probably not relevant for genetic selection against mastitis in the analyzed breed.

Screening of Three Different Alleles of mtDNA (G709A, G3496T, A3537G) in Subpopulation of UKM Students

Human DNA consists of nucleus DNA (nDNA) and mitochondrial DNA (mtDNA). Both are valuable in medicine and forensic genetics but in this project, single nucleotide polymorphisms (SNPs) in mtDNA are used to trace the mutation occurred. Mutations in the sequence of alleles can lead to haplogroup variation and also certain diseases. The purpose of this study is to screen of mutations on alleles G709A, G3496T, and A3537G in Malay population of The National University of Malaysia (UKM) students. These SNPs lie in the ND1 (nitrogen dehydrogenase subunit 1) coding region, and the reports state that these three alleles are prone to mutate. From MitoMap Web site, the mutations of these alleles are reported to have potential in causing several diseases with the collaboration of other SNPs mutation. Allele G709A is reported to have an association with hearing loss and Leber Hereditary Optic Neuropathy (LHON) while allele G3496T is associated to LHON only. Allele A3537G is related to diabetes. A total of 100 DNA samples were collected from Malay students of UKM and preserved on FTA card to be purified later. The concentration of the DNA on the purified FTA card was between 10μM to 20μM. An attempt was made by amplifying those three loci from the genomic DNA. The amplified product was detected and separated using 1% gel electrophoresis. Before sequencing, the PCR products were visualized under UV light using gel documentation system. All PCR products were sequenced to detect the mutation on every single position chosen. From the alignment of sequencing results, allele G709A and allele G3496T showed no mutation. Meanwhile four samples from alleles A3537G has the mutation. From the results obtained, it seems that mutations are rare in all selected alleles. It is recommended to increase the sample size and alleles selected in the future to increase the strength of the study. This study also should be applied to other populations in Malaysia such as Chinese and Indian.&#x0D;

L-selectin gene polymorphism and its association with clinical mastitis, somatic cell score, and milk production in Polish Holstein-Friesian cattle

Bovine mastitis is a widespread disease of the mammary gland, highly contributing to the increase in veterinary costs in dairy industry. In the present study, the genetic polymorphism within bovine L-selectin gene was analysed and its impact on clinical mastitis occurrence, somatic cell score (SCS), and milk production traits in Polish Holstein-Friesian cows was examined. Polymorphism within L-selectin gene, molecule responsible for neutrophil attachment to endothelium, might have a potential role in immune response to bacterial infections and udder health. Two hundred and six Polish Holstein-Friesian cows were genotyped by polymerase chain reaction-restriction fragment length polymorphism method. Two single nucleotide polymorphisms mutations within the coding sequence of L-selectin gene were identified (c.165G&amp;gt;A and c.567C&amp;gt;T). The effect of c.165G&amp;gt;A and c.567C&amp;gt;T mutations on SCS was highly significant (P = 0.0019 and P = 0.0003, respectively). Strong associations (P ≤ 0.0001) were also observed between L-selectin polymorphism and milk production traits (milk yield, milk fat percentage, and milk protein percentage). However, the polymorphism in the analysed gene had no influence on the resistance or susceptibility of cows to clinical mastitis (only the tendency toward significance, P = 0.06 for c.567C&amp;gt;T mutation was found). Potential exploitation of the information on the identified associations in genetic selection needs to confirm the obtained results in further investigations.

Identification of the dwarf gene GmDW1 in soybean (Glycine max L.) by combining mapping-by-sequencing and linkage analysis

Key messageGmDW1 encodes an ent-kaurene synthase (KS) acting at the early step of the biosynthesis pathway for gibberellins (GAs) and regulates the development of plant height in soybean.Plant height is an important component of plant architecture, and significantly affects crop breeding practices and yield. Here, we report the characterization of an EMS-induced dwarf mutant (dw) of the soybean cultivar Zhongpin 661 (ZDD23893). The dw mutant displayed reduced plant height and shortened internodes, both of which were mainly attributed to the longitudinally decreased cell length. The bioactive GA1 (gibberellin A1) and GA4 (gibberellin A4) were not detectable in the stem of dw, and the dwarf phenotype could be rescued by treatment with exogenous GA3. Genetic analysis showed that the dwarf trait of dw was controlled by a recessive nuclear gene. By combining linkage analysis and mapping-by-sequencing, we mapped the GmDW1 gene to an approximately 460-kb region on chromosome (Chr.) 8, containing 36 annotated genes in the reference Willliams 82 genome. Of these genes, we identified two nonsynonymous single nucleotide polymorphisms (SNPs) that are present in the encoding regions of Gmdw1 and Glyma.08G165100 in dw, respectively. However, only the SNP mutation (T>A) at nucleotide 1224 in Gmdw1 cosegregated with the dwarf phenotype. GmDW1 encodes an ent-kaurene synthase, and was expressed in various tissues including root, stem, and leaf. Further phenotypic analysis of the allelic variations in soybean accessions strongly indicated that GmDW1 is responsible for the dwarf phenotype in dw. Our results provide important information for improving our understanding of the genetics of soybean plant height and crop breeding.

Single Nucleotide Polymorphisms (SNPs) of URAT1 (rs7932775) and ABCG2 (rs3825016) on Chronic Kidney Disease Patients with Hyperuricemia

Background: More and more chronic kidney disease (CKD) patients are accompanied with hyperuricaemia. As is known, hyperuricaemia is an independent hazard of both cardiovascular diseases (CVD) and chronic kidney diseases. We aim at identifying Single Nucleotide Polymorphism (SNP) difference of hURAT1 (rs7932775) and ABCG2 (rs3825016) on CKD patient with hyperuricemia and/or gout. Methods: All forty-two CKD patients were divided into two groups: hyperuricemia, and control group. 24 hours urine sample and serum were prepared for testing biochemistry parameters. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method is used to analyze hURAT1 and ABCG2 single nucleotide polymorphisms in different groups. Results: 17 patients have CT SNP of hURAT1 (rs7932775) and 13 patients have CT SNP of ABCG2 (rs3825016) in hyperuricemia group, while only 5 persons and 6 persons have the same mutations in control group respectively. 7 patients have CT SNP of both hURAT1 (rs7932775) and ABCG2 (rs3825016) in hyperuricemia group, while only 2 persons have the same mutations in control group. CT mutation rates of hURAT1 (rs7932775) and ABCG2 (rs3825016) in hyperuricemia group were 60.7% (17/28) and 50% (13/28) respectively, higher than that of control group (35.7% (5/14) and 42.8% (6/14)). What is more, Double SNP mutations in both hURAT1 (rs7932775) and ABCG2 (rs3825016) in hyperuricemia group were 25% (7/28), higher than that of control group (14.2%, 2/14). Conclusion: There are higher mutation rates of CT SNP in hURAT1 (rs7932775) and/or ABCG2 (rs3825016) in hyperuricemia group. We can conclude that hyperuricemia is a high risk factor in progress of CKD, which is necessary to take measures of decreasing serum uric acid to delay CKD progress.