Study on the Molecular Mechanism of Cadmium Uptake and Transport Mediated by Chuanxiong ZIP Family Genes
Description
This dataset comprises the raw data from the study investigating the molecular mechanisms of cadmium uptake and transport mediated by ZIP family genes in Ligusticum chuanxiong
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Steps to reproduce
Following harvest of the entire Ligusticum chuanxiong plant, it was immersed in 10 mmol·L⁻¹ EDTA-Na₂ for 20 minutes to remove surface-adsorbed Cd²⁺. The plant was then separated into four parts: rhizome, aerial stem, leaves, and leaf veins, and dried in a 60°C oven to constant weight. Accurately weigh 0.5 g of dried sample, digest using nitric acid, and determine the content of each metallic element by inductively coupled plasma mass spectrometry (ICP-MS). The same method was employed for determining Cd content in Arabidopsis thaliana.Control group rhizomes and aerial parts stored at -80°C, alongside those treated with 200 μmol·L⁻¹ CdCl₂ for 14 days, were subjected to total RNA extraction using the TRIzol method. Following quality control verification, mRNA was enriched via the NEBNext Poly(A) mRNA Magnetic Isolation Module, and sequencing libraries were constructed. Sequencing was performed using the Illumina HiSeq 2500 platform (Beijing Novogene Co., Ltd.). Raw data underwent quality control via fastp (v0.20.0) to obtain clean data . Gene expression levels (TPM values) were calculated using RSEM (v1.3.3), and differentially expressed genes were identified via DESeq2 (v1.24.0) based on criteria of Fold Change ≥ 1.5 and adjusted P-value < 0.05.To determine quantitative fluorescence data, RNA was extracted from liquid nitrogen-stored control group and 200 μmol·L⁻¹ CdCl₂ treated Ligusticum chuanxiong rhizome and aerial stem samples at 1, 3, 7, and 14 days. Following verification of RNA concentration and purity, cDNA was synthesised using the PrimeScript™ RT Kit (Takara, Japan). Using Actin as the internal reference gene, qRT-PCR analysis was performed with the TB Green® Premix Ex Taq™ II (Tli RNaseH Plus) kit (Takara, Japan) on a Bio-Rad CFX96 real-time fluorescent quantitative PCR instrument (Bio-Rad, USA), employing a 20 μL reaction system. Relative expression levels of LcZIPs genes were calculated using the 2⁻ΔΔCT method, with three biological replicates per sample.Moreover,Lithium acetate-mediated transformation was employed to introduce the empty vector and the pAG413-LcZIPs yeast expression vector into the cadmium-sensitive yeast mutant Δycf1. A single colony of the transgenic yeast strain was cultured in SD-his-glu liquid medium with shaking until an OD600 of 1 was reached. The cell suspension was then serially diluted at 10, 100, 1000, and 10000-fold dilutions. Ten microlitres of cell suspension were sequentially spotted onto control and SD solid media containing 50 μM CdCl₂, incubated statically at 30°C for 2–3 days, and photographed for analysis. To further assess the growth of transgenic yeast under cadmium or zinc stress, the strain was inoculated into SD-his-gal liquid medium containing CdCl₂ (0, 15 μM, 25 μM). OD₆₀₀ values were recorded at 3 h, 6 h, 9 h, 12 h, 24 h, 32 h, and 35 h.
Institutions
- Tianjin University of Traditional Chinese Medicine