Disruption of a DUF247 Containing Protein Alters Cell Wall Polysaccharides and Reduces Growth in Arabidopsis.

Plant cell wall biosynthesis is a complex process that requires proteins and enzymes from glycan synthesis to wall assembly. We show that disruption of At3g50120 (DUF247-1), a member of the DUF247 multigene family containing 28 genes in Arabidopsis, results in alterations to the structure and composition of cell wall polysaccharides and reduced growth and plant size. An ELISA using cell wall antibodies shows that the mutants also exhibit ~50% reductions in xyloglucan (XyG), glucuronoxylan (GX) and heteromannan (HM) epitopes in the NaOH fraction and ~50% increases in homogalacturonan (HG) epitopes in the CDTA fraction. Furthermore, the polymer sizes of XyGs and GXs are reduced with concomitant increases in short-chain polymers, while those of HGs and mHGs are slightly increased. Complementation using 35S:DUF247-1 partially recovers the XyG and HG content, but not those of GX and HM, suggesting that DUF247-1 is more closely associated with XyGs and HGs. DUF247-1 is expressed throughout Arabidopsis, particularly in vascular and developing tissues, and its disruption affects the expression of other gene members, indicating a regulatory control role within the gene family. Our results demonstrate that DUF247-1 is required for normal cell wall composition and structure and Arabidopsis growth.

Gel purification of gDNA for next-generation sequencing applications.

We demonstrate that gDNA can be conveniently and efficiently isolated and purified using standard agarose gel electrophoresis, band excision and gel purification. This method yields a substantial amount at microgram levels of gDNA per gel cleanup with high purity. An RNase A treatment step can be omitted. The quality of gDNA is suitable for next-generation sequencing, resulting in >10 Mb reads and high-quality read data (Phred score >28 up to 100 of 150 base reads). Furthermore, the gDNA can be kept intact in a gel slice for several days. This method has been tested for dictyostelids, bacteria and plants.

Evaluations of the mutagenicity of a pigment extract from bulb culture of Hippeastrum reticulatum.

The use of anthocyanins in food products as colorants has been limited because of their instability toward alkaline pH and high temperature. This study aimed to determine color stability and mutagenicity of the anthocyanin-based pigment extract from bulb cultures of Hippeastrum (Hippeastrum reticulatum). The pigment extract retained its reddish-orange color under alkaline conditions (⩽pH 11) and was stable up to 6 h at 95 °C. The mutagenicity of the extract was evaluated in vitro and in vivo. Hippeastrum pigment extract up to 1.25 mg plate(-1) was found non-mutagenic in Ames test using Salmonella typhimurium strain TA98 and TA100. Chromosome aberrations were observed when human lymphocytes were treated with the extract up to 1.5 mg ml(-1). However, the extract up to 1.4 mg ml(-1) was found to exhibit relatively low or no mutagenicity in in vitro comet assays with human lymphocytes. In in vivo micronucleated reticulocyte assay, mice were treated orally with the extract up to 1 g kg(-1). No significant increase of the percentage of micronucleated peripheral reticulocytes compared to the negative control groups was found. Taken together, our study indicates that Hippeastrum pigment extract is potentially applicable as an additive colorant in the diet and related products.