Knowledge of the Se fractionation and the role of dissolved organic matter (DOM) in soil is the key to understanding Se mobility and its bioavailability in the soil-plant system. In this study, single extractions using phosphate-buffer (PBS), sequential extraction procedures (SEP), and diffusive gradients in thin-films (DGT) were used to measure Se bioavailability in soil supplemented with selenite and organic amendment (cow and chicken manures). Selenium fraction was isolated into DOM-Se fractions, such as hydrophilic acid-bound Se (HY-Se), fulvic acid-bound Se (FA-Se), humic acid-bound Se (HA-Se), and hydrophobic organic neutral-bound Se (HON-Se), by a rapid batch technique using XAD-8 resin (AMBERLITE XAD™, USA). Simultaneous application of either cow or chicken manure with selenite could result in the decrease of Se availability in the soil. Isolating Se available fraction into DOM-Se fractions showed that low-molecular-weight DOM-Se as an available fraction and even HY-Se as a less available fraction (OM-Se) were likely the major sources for Brassica juncea (L.) Czern. et Coss uptake in soil. Moreover, knowledge of the DOM-Se composition, especially the low-molecular-weight DOM-Se fractions, is important for assessing the bioavailability of Se in soil, the results of which are more accurate than the chemical extraction method. The high value of Pearson correlation coefficients between CDGT-Se and Se concentrations in shoots, tubers and roots of Brassica juncea (L.) Czern. et Coss in cow and chicken manures treatment were 0.95 and 0.99, 0.96 and 0,96, and 0.89 and 0.97 (p < 0,05), respectively, indicating that DGT-Se can reflect the Se uptake ability by plants and can be used to predict the bioavailability of Se when manure and selenite are simultaneously applied. Unlabelled Image • DGT is a reliable method for evaluating Se bioavailability in organic enrich soils. • Not all Se species in PBS- and SEP-labile fraction can be taken up by plants. • Isolation of the Se fraction into DOM-Se fractions can improve Se bioavailability assessments • DOM play dual effect of on fraction transformation and availability of Se in soils. [ABSTRACT FROM AUTHOR]