Gene improving cadmium tolerance of plant and application thereof
A cadmium, plant-resistant technology, applied in the field of bioengineering, can solve problems such as damage to the normal function of the body, soil, water source damage, etc.
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Embodiment 1
[0033] Embodiment 1, the acquisition of XCD1 and its coding gene
[0034] Using chemical induction to activate the XVE (LexA-VP16-ER) mutant system (provided by the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences), from containing 100 μM CdCl 2 The Arabidopsis seedlings screened in the 1 / 2MS medium were used as materials ( figure 1 ), use the mutant genomic DNA extracted by the classic CTAB method as a template, and perform the following Tail-PCR reaction (Yao-Guang Liu et al., 1995):
[0035] Tail-PCR reaction program (including three rounds of reaction, Table 1):
[0036]
[0037]
[0038] Table 1 Tail-PCR reaction program
[0039]
[0040] Among them, nested primers: LexA2, LexA3, LexA4, LexA5, LexA6, and random primers: AD1, AD2, AD3, AD4 are shown in Table 2.
[0041] Table 2 Primer sequences used in Tail-PCR
[0042]
[0043]
[0044] Perform 1% agarose gel electrophoresis on the third-round product of Tail-PCR. The obtained ...
Embodiment 2
[0045] Embodiment 2, cultivate cadmium-tolerant Arabidopsis thaliana
[0046] 1. Obtaining and phenotypic analysis of cadmium-tolerant mutant xcd1
[0047] The Arabidopsis mutant library of independent transformation lines constructed by chemically inducing and activating the XVE (LexA-VP16-ER) mutant system, under the stress condition of 100 μM CdCl2 and the induction of 10 μM external hormone β-estradiol, at 1 / The WT and mutants were sowed in 2MS medium and cultured horizontally for 1w, and several cadmium-tolerant mutants were screened ( figure 1 ), the larger seedlings were selected for follow-up research, and the gain-of-function cadmium-tolerant mutant xcd1 was obtained through phenotypic identification (Fig. 2A-C).
[0048] 2. Comparison of cadmium accumulation between xcd1 and wild-type plants
[0049] Since the induction and activation of XCD1 will significantly enhance the plant's tolerance to heavy metal Cd, whether the Cd content in the mutant also changes. Ther...
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