Magnetic Bead Adsorption Extraction of Xyloglucan Endoglucosidase/Hydrolase Gene and Its Expression Analysis in Land Cotton

Abstract

The xyloglucan Endotransglucosylase/hydrolase (XTH) genes are proposed to encode enzymes responsible for cleaving and reattaching xyloglucan polymers. Despite prior identification of the XTH gene family in Arabidopsis and rice, the XTH family in upland cotton, a tetraploid plant whose fiber cell is an excellent model for the study of plant cell elongation, is yet uncharacterized. In this study, iron tetroxide based magnetic nanobead (Fe3O4 NPs) was successfully prepared and applied to extract xyloglucan endoglucosidase/hydrolase genes. Analysis of the genes can provide insight into the evolutionary significance and function of the XTH gene family. A total of 41 XTH genes found by searching the phytozomev 10 database were classified into three groups based on their phylogeny and the motifs of individual genes. The 25 and 5 GhXTH genes occurred as clusters resulting from the segmental and tandem duplication. More frequent duplication events in cotton contributed to the expansion of the family. Global microarray analysis of GhXTH gene expression in cotton fibers showed that 18 GhXTH genes could be divided into two clusters and four subclusters based on their expression patterns. Accumulated expression levels were relatively high at the elongation stages of the cotton fibers, suggesting that cotton fiber elongation requires high amounts of the GhXTH protein. The expression profiles of GhXTH3 and GhXTH4 showed by quantitative realtime PCR were similar to those determined by microarray. Additionally, the expression levels of GhXTH3 and GhXTH4 in Gossypium barbadense were higher than those in Gossypium hirsutum at developmental stages, indicating that expression levels of GhXTH3 and GhXTH4 in fibers varied among cultivars differing in fiber length.