Genomic exploration and molecular marker development in a large and complex conifer genome using RADseq and mRNAseq.
Mol Ecol Resour. 2014 Sep 15;
Authors: Karam MJ, Lefèvre F, Dagher-Kharrat MB, Pinosio S, Vendramin GG
We combined Restriction site Associated DNA sequencing (RADseq) using a hypomethylation-sensitive enzyme and messenger RNA sequencing (mRNAseq) to develop molecular markers for the 16 Gigabase genome of Cedrus atlantica, a conifer tree species. With each method, Illumina(®) reads from one individual were used to generate de novo assemblies. SNPs from the RADseq dataset were detected in a panel of one single individual and three pools of three individuals each. We developed a flexible script to estimate the ascertainment bias in SNP detection considering the pooling and sampling effects on the probability of not detecting an existing polymorphism. Gene Ontology (GO) and Transposable Element (TE) search analyses were applied to both datasets. The RADseq and the mRNAseq assemblies represented 0.1% and 0.6% of the genome, respectively. Genome complexity reduction resulted in 17% of the RADseq contigs potentially coding for proteins. This rate was doubled in the mRNAseq dataset, suggesting that RADseq also explores non-coding low repeat regions. The two methods gave very similar GO-slim profiles. As expected, the two assemblies were poor in TE-like sequences (<4% of contigs length). We identified 17,348 single nucleotide polymorphisms (SNPs) in the RADseq dataset and 5,714 simple sequence repeats (SSRs) in the transcriptome. A subset of 282 SNPs was validated using the Fluidigm genotyping technology, giving a conversion rate of 50.4%, falling within the expected range for conifers. Increasing sample size had the greatest effect for ascertainment bias reduction. These results validated the utility of the RADseq approach for highly complex genomes such as conifers. This article is protected by copyright. All rights reserved.
PMID: 25224750 [PubMed - as supplied by publisher]
Molecular genetic analysis and structure model of a rare B(A)02 subgroup of the ABO blood group system.
Transfus Apher Sci. 2014 Aug 27;
Authors: Chen Q, Li J, Xiao J, Du L, Li M, Yao G
BACKGROUND: Serological analysis of ABO blood group has been widely applied in transfusion medicine. However, ABO subgroups with different expression of blood group antigens sometimes cannot be determined by serological methods. Therefore, genotyping is useful to understand the variant ABO phenotypes.
MATERIAL AND METHODS: Exon 6 to exon 7 and adjacent introns of the ABO gene from a donor with ABO typing discrepancy were amplified and sequenced. Cloning sequencing was also performed to identify the allele. To explore the effect of mutation, three dimensional model of mutant p.Pro234Ala was built and optimized.
RESULTS: The variant B (c. 700C > G) allele expressed an AweakB phenotype with anti-A in his serum with a ABO*B(A)02/O02 heterozygote genotype. Cloning sequencing confirmed that the c.700C > G single nucleotide polymorphism was associated with a B101 allele. Three dimensional molecular modeling suggested that p.Pro234Ala might affect the conformation of His233, Met266 and Ala268, which were known as critical residues for donor recognition.
CONCLUSION: ABO genotyping is needed for correct identification subgroups to improve accuracy evaluation of blood typing and increase the safety of blood transfusion. Alteration of DNA sequence in the ABO gene resulted in amino acid substitutions and led to a weak or missing expression of ABO antigens.
PMID: 25217989 [PubMed - as supplied by publisher]
GWAS of 972 autologous stem cell recipients with multiple myeloma identifies 11 genetic variants associated with chemotherapy-induced oral mucositis.
Support Care Cancer. 2014 Sep 14;
Authors: Coleman EA, Lee JY, Erickson SW, Goodwin JA, Sanathkumar N, Raj VR, Zhou D, McKelvey KD, Apewokin S, Stephens O, Enderlin CA, Vangsted AJ, Reed PJ, Anaissie EJ
PURPOSE: High-dose chemotherapy and autologous stem cell transplant (ASCT) to treat multiple myeloma (MM) and other cancers carries the risk of oral mucositis (OM) with sequelae including impaired nutritional and fluid intake, pain, and infectious complications. As a result of these problems, cancer treatment may have to be interrupted or delayed. In this study, we looked beyond OM's known risk factors of renal function and melphalan dose with a genome-wide association study (GWAS) to evaluate whether genetic variants in conjunction with clinical risk factors influence predisposition for OM.
METHODS: Genotyping was performed using Illumina HumanOmni1-Quad v1.0 BeadChip and further assessed for data quality. We tested 892,589 germline single-nucleotide polymorphisms (SNPs) for association with OM among 972 Caucasian patients treated with high-dose melphalan and ASCT in Total Therapy clinical trials (TT2, TT3, TT4) for newly diagnosed MM. Statistical analyses included t tests, stepwise regression modeling, and logistic regression modeling to find baseline clinical factors and genotypes associated with OM.
RESULTS: We found that 353 (36.3 %) patients had grades 2-4 OM. Type of treatment protocol, baseline estimated glomerular filtration rate, and melphalan dose along with baseline serum albumin and female gender predicted 43.6 % of grades 2-4 OM cases. Eleven SNPs located in or near matrix metalloproteinase 13, JPH3, DHRS7C, CEP192, CPEB1/LINC00692, FBN2, ALDH1A1, and DMRTA1/FLJ35282 were associated with grades 2-4 OM. The addition of these SNPs increased sensitivity in detecting grades 2-4 OM cases to 52 %.
CONCLUSIONS: These SNPs may be important for their roles in inflammatory pathways, epithelial healing, and chemotherapy detoxification.
PMID: 25218607 [PubMed - as supplied by publisher]
Rare allele of HvLox-1 associated with lipoxygenase activity in barley (Hordeum vulgare L.).
Theor Appl Genet. 2014 Sep 12;
Authors: Guo G, Dondup D, Yuan X, Gu F, Wang D, Jia F, Lin Z, Baum M, Zhang J
KEY MESSAGE: Identification and allele-specific marker development of a functional SNP of HvLox - 1 which associated with barley lipoxygenase activity. Improving the stability of the flavor of beer is one of the main objectives in breeding barley for malting, and lipoxygenase-1 (LOX-1) is a key enzyme controlling this trait. In this study, a modified LOX activity assay was used for null LOX-1 mutant screening. Four barley landraces with no detected level of LOX-1 activity were screened from 1,083 barley germplasm accessions from China. The genomic sequence diversity of the HvLox-1 gene of the four null LOX-1 Chinese landraces was compared with that of a further 76 accessions. A total of 104 nucleotide polymorphisms were found, which contained 83 single-nucleotide polymorphisms (SNPs), 7 multiple-nucleotide polymorphisms, and 14 insertions and deletions. Most notably, we found a rare C/G mutation (SNP-61) in the second intron which led to null LOX-1 activity through an altered splicing acceptor site. In addition, an allele-specific polymerase chain reaction marker was developed for the genotyping of SNP-61, which could be used in breeding programs for barley to be used for malting. The objective was to improve beer quality.
PMID: 25212109 [PubMed - as supplied by publisher]