Genome-wide Association Analysis Of Vitamin B In Common Wheat

Vitamin B,one of the microelements,is essential for maintaining normal life activities.Instead of synthesizing by themselves,human and animals have to intake Vitamin B from outside world,especially from cereal products which is one of the most important staple food sources.Therefore,the research of Vitamin B content in cereal is significant for improving wheat quality and human health.The objective of this study was to develop an optimal procedure for extraction and establish a valid method for determination of Vitamin B1,B2,and B6 using high performance liquid chromatograph(HPLC).The content of Vitamin B1 and B2 in 166 wheat cultivars were genotyped with wheat 90 K iSelect chip.The main results are summarized as follows:1.Vitamin B from bread wheat cultivars were extracted using solvent extraction based on the report as described by previous research for color wheat with modification.The highest extraction yield of Vitamin B1 and B2 was achieved in 0.05 M sodium acetate(pH 4.5)followed by the addition of papain,1% glutathione,acid phosphatase and α-amylase.The extraction of Vitamin B6 was extracted in a solution of the above extraction adding glyoxylic acid and ferrous sulfate.The separation was achieved using Waters XTerra RP18(150×4.6 mm,5 μm)with RF10 Axl fluorescence detector,and the column was thermostated at 35°C.The temperature of the sample chamber was 4℃ with equal elution.Under a gradient system consisting of methanol(A)and 0.05 M sodium acetate solution pH 4.5(B)(30:70,v/v)at a constant flow rate of 0.7 ml/min for Vitamin B1 and B2.The detector operated at an excitation/emission wavelength of 366 nm/435 nm for Vitamin B1,and at an excitation/emission wavelength of 422 nm/522 nm for Vitamin B2.For Vitamin B6,the gradient system consisting of methanol(A)and 0.05 M potassium dihydrogen phosphate solution containing 5 mM sodium heptanesulfonate(B)(3:97,v/v)at a constant flow rate of 0.7 ml/min.The B mobile phase was then adjusted to pH 2.5 using phosphoric acid.The detector operated at an excitation / emission wavelength of 290 nm/395 nm for Vitamin B6.This method should be easier and suitable for determining lots of samples.2.A genome-wide association study(GWAS)consisted of 18,207 SNP markers was performed with a panel of 166 wheat cultivars.Twenty-four loci were identified.Among these marker-trait associations(MTAs),17 loci were significantly identified for Vitamin B1 content and explained 6.5-10.6% of phenotypic variances.IWB43809(6AS,0cM)and IWB69903(6AS,13cM)were detected simultaneously in two environments.A total of 7 MTAs were identified for Vitamin B2 content and explained 6.5-10.5% of phenotypic variances.These results could provide a basis for improving Vitamin B1 and B2 content in common wheat.