激发光源用于花青素的生物合成的研究

近日，福建省农业科学院食品科学与技术研究所发表文献《Whole-pathway stimulation of anthocyanin biosynthesis by UDP-glycose: flavonoid glycosyltransferase gene (UFGT) providing new insights into natural product development based on plant cell factory》，文献中实验使用了LUYOR-3415RG便携式双波长荧光蛋白激发光源用于观察GFP在愈伤中的表达，用于筛选转基因阳性的细胞。

文献摘要:

Detection of positive transformed cell lines

The resistance cell lines were observed using a fluorescent protein hand-held lamp (LUYOR-3415, Shanghai, China). The cell lines exhibiting green fluorescence characteristics were preliminary identified as positive transformed cell lines. These positive cell lines underwent further screening for three generations, with decreasing resistant concentrations, and were subsequently transferred to solid MS supplemented with 1.0 mg/L 2, 4-D without antibiotics for further subculture. Genomic DNA was extracted for PCR and sequencing to verify the transformation. And subcellular localization observation of UFGT was conducted using a FV1200 confocal microscope (Olympus, Tokyo, Japan).

3.2. Characteristics of transgenic cells and screening of high-yield anthocyanin cell line

Transgenic screening identified four positive grape cell lines (Fig. 2A), which exhibited a green fluorescent signal that diminished or disappeared with extended culture duration due to cellular senescence. Therefore, a portable fluorescent excitation light facilitates comprehensive full-stage observation and screening of transformed cells in vivo. These four positive grape cell lines were further validated using DNA-PCR (Fig. 2B). Subcellular localization analysis revealed that VdUFGT was localized in both the cytoplasm and nucleus (Fig. 2C), suggesting its potential role in the process. Both the WT and four VdUFGT transgenic cell lines exhibited varying degrees of red color (Fig. 2D). The transgenic cell lines generally displayed a deep red color on the surface, while the WT appeared light red, indicating a higher accumulation of anthocyanin in the transgenic cell lines. The proliferation coefficient results showed no significant differences among the different cell lines (Fig. 2E), suggesting that the overexpression of VdUFGT did not have a significant impact on the growth of grape cells. The anthocyanin content in the overexpressed cell lines was significantly higher than that in WT cell line, with the vgt3 cell line reaching 195.2 μg/g (FW), a 4.2-fold increase compared to the WT (Fig. 2F). However, the flavonoid content in vgt3 was significantly lower than that in WT cell line (1504.6 μg/g FW) (Fig. 2G). This discrepancy may be attributed to anthocyanins being terminal metabolites while flavonoids are intermediate ones. Consequently, the overexpression of VdUFGT promoted anthocyanin accumulation, with the vgt3 identified as a high-yield cell line for further utilization.

文献地址：https://doi.org/10.1016/j.fbio.2024.105409 

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