Arabidopsis thaliana stress-resistance related gene AtZAT6, preparation method and application
An Arabidopsis thaliana and gene technology, applied in the preparation of Arabidopsis thaliana stress-related gene AtZAT6, the AtZAT6 gene that enhances plant resistance to drought and salt stress, and the Arabidopsis stress-related gene AtZAT6 in enhancing plant resistance to drought and In the field of salt stress resistance, it can solve problems such as crop yield reduction and natural ecosystem degradation
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Embodiment 1
[0042] an Arabidopsis AtZAT6 A gene preparation method, the steps of which are:
[0043] A. Grind young plant leaf samples into powder in liquid nitrogen, and use the Trirol extraction kit (purchased from Invitrogen, the same below) to extract RNA. The extracted total RNA was dissolved in RNase-free double distilled water. DNase I (purchased from Promega, the same below) removed possible residual DNA. A protein detector (DM 650 BECKMAN, MSA) was used to detect the light absorbance of RNA at 260 nm and 280 nm, respectively, and combined with 1% (mass volume ratio) agarose gel electrophoresis to identify the purity and concentration of RNA. The obtained RNA was used as a template for reverse transcription, and the obtained cDNA was aliquoted and stored at -20°C for future use.
[0044] B. According to the Arabidopsis public database TAIR (http: / / www.arabidopsis.org / ) in Arabidopsis AtZAT6 Gene sequence (At5g043400), primers were designed in the gene CDS (Coding seqμence, cod...
Embodiment 2
[0046] A resistance-related gene in Arabidopsis thaliana AtZAT6 Obtaining of overexpression plants:
[0047] 1, AtZAT6 Gene overexpression vector (OX- AtZAT6 ) construction and transformation of Agrobacterium tumefaciens strain GV3101:
[0048] The constructed recombinant vector is the constructed plasmid pBIM (preserved in our laboratory, Yang LX, Wang RY, Ren F, Liu J, Cheng J, Lu YT (2005) AtGLB1 enhances the tolerance of Arabidopsis to hydrogen peroxide stress. Plant Cell Physiol 46: 1309-1316. See Genetic Resources Table for details) The target gene on the AtZAT6 Gene replacement. To accomplish this, first use the SmaI / XhoI double digest cloned AtZAT6 gene fragments, while using SmaI / XhoI Digested pBIM plasmid (preserved in our laboratory, Yang LX, Wang RY, Ren F, Liu J, Cheng J, Lu YT (2005) AtGLB1 enhances the tolerance of Arabidopsis to hydrogen peroxide stress. Plant Cell Physiol 46: 1309-1316. See Genetic Resources Table for details). The enzyme ...
Embodiment 3
[0057] a resistance-related AtZAT6 The application of genes in enhancing the resistance of plants (Arabidopsis thaliana) to drought and salt stress, the steps are:
[0058] A. The Arabidopsis OX- AtZAT6 Overexpression strains and wild-type seeds were vernalized at 4°C for one week, and then sowed in a light culture room at 23°C for moisturizing culture (16 h of light and 8 h of darkness per day) for 14 days, and the nutrient solution was poured every 3 days within 14 days (the composition of the nutrient solution For: 5ml per liter of 1 M KNO 3 , 2 ml 1 M Ca (NO 3 ) 2˙ 4H 2 O, 2 ml 1M MgSO 4˙ 7 h 2 O, 2.5 ml 20 mM Fe ˙ EDTA, 2.5 ml 1 M phosphate buffer (pH5.5), 1 ml MS trace elements);
[0059] B. Select 14-day Arabidopsis seedlings (including OX- AtZAT6 Overexpression strain and wild type) were divided into three groups on average for 21 days of experiment, the specific experiment is as follows:
[0060] 1), the first group of Arabidopsis seedlings (including OX- A...
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