Development and validation of an SNP genotyping array and construction of a high-density linkage map in castor.

Castor is a commercially important oilseed crop that provides raw materials for several industries. Currently, the availability of genomic resources for castor is very limited. In this study, genome-wide SNPs were discovered in castor via whole-genome sequencing of 14 diverse lines to an average of 34X coverage. A total of 2,179,759 putative SNPs were detected, and a genotyping array was designed with 6,000 high-quality SNPs representing 2,492 scaffolds of the draft castor genome (87.5% genome coverage). The array was validated by genotyping a panel of 314 inbred castor lines, which resulted in 5,025 scorable SNPs with a high call rate (98%) and reproducibility (100%). Using this array, a consensus linkage map consisting of 1,978 SNP loci was constructed with an average inter-marker distance of 0.55 cM. The genome-wide SNP data, the genotyping array and the dense linkage map are valuable genomic tools for promoting high-throughput genomic research and molecular breeding in castor.

Validation of a method for diosgenin extraction from fenugreek (Trigonella foenum-graecum L.).

BACKGROUND: Diosgenin is a very important plant secondary metabolite and raw material for the drug indus- try. Plant sources rich in diosgenin include yam (Dioscorea spp.) and fenugreek (Trigonella foenum-graecum L.). A method for diosgenin extraction from yam extracts has previously been validated, but its extraction from fenugreek plants still requires validation. In addition, all available methods require time-consuming additional purification steps. The present study was aimed at developing a low cost, less time-consuming single-step method for diosgenin extraction from fenugreek. METHODS: This study represents a method developed for diosgenin extraction from fenugreek plants without any additional/supportive purification methods such as chromatography or thin-layer chroma- tography. Diosgenin yield estimation and purity analysis by HPLC method, along with accuracy and preci- sion analysis, is presented. RESULTS: Five different fenugreek varieties were subjected to a newly developed diosgenin extraction method, and an HPLC chromatogram showed a single peak corresponding to diosgenin. Yield was determined by the standard curve method. Limit of detection (LOD) and limit of quantification (LOQ) for the assay were found to be 0.0312 and 0.102 μg, respectively; tcalculated for slope and other statistical parameters were found to be significant (P value < 0.001) for this method. CONCLUSIONS: We have developed a fast, accurate and low cost method for diosgenin extraction from fenu- greek. Although the authors have studied this method only in fenugreek plants, it could be applied to the extraction of a few other plant secondary metabolites, which will help researchers to save time and effort.

Compound heterozygous ß(+) ß(0) mutation of HBB gene leading to ß-thalassemia major in a Gujarati family - A case study.

ß-Thalassemia is a genetic disease characterized by reduced or non-functionality of ß-globin gene expression, which is caused due to a number of variations and indels (insertions and deletions). In this case study, we have reported a rare occurrence of compound heterozygosity of two different variants, namely, HBBc.92G > C and HBBc.92 + 5G > C in maternal amniotic fluid sample. Prenatal ß-thalassemia mutation detection in fetal DNA was carried out using nucleotide sequencing method. After analysis, the father was found to be heterozygous for HBBc.92G > C (Codon 30 (G > C)) mutation which is ß(0) type and the mother was heterozygous for HBBc.92 + 5G > C (IVS I-5 (G > C)) mutation which is ß(+) type. When amniotic fluid sample was analyzed for ß-globin gene (HBB), we found the occurrence of heterozygous allelic pattern for aforesaid mutations. This compound heterozygous state of fetus sample was considered as ß(+)/ß(0) category of ß thalassemia which was clinically and genotypically interpreted as ß-thalassemia major. Regular blood transfusions are required for the survival of thalassemia major patients hence prenatal diagnosis is imperative for timely patient management. Prenatal diagnosis helps the parents to know the thalassemic status of the fetus and enables an early decision on the pregnancy. In the present study, we have identified compound heterozygosity for ß-thalassemia in the fetus which portrays the importance of prenatal screening.