Application of dap gene in Deinococcus radiodurans R1 in breeding of salt tolerant plants
A gene and plant technology, applied in the field of application of the gene in improving plant resistance to salt stress, can solve unseen problems
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Embodiment 2
[0037] Example 2 Salt Tolerance Experiment Containing D.radiodurans R1dap Gene (DRB0118) Recombinant Strain
[0038] 1. Experimental method
[0039] 1. The 2 recombinant Escherichia coli obtained in Example 1 were respectively inoculated in 20 mL of LB liquid medium (containing Amp antibiotics), and after the shake flask was cultured overnight (37° C.), they were then respectively transferred to 100 mL of LB liquid medium In the medium, try to keep the inoculum volume consistent, culture to OD 600 ≈0.5 is enough.
[0040] 2. After centrifuging 10mL of the bacterial solution, shock it in an equal volume of 4M NaCl salt solution for 2 hours, and immediately dilute each sample to 10 times with sterile deionized water. -4 , Take 10 μL and spot on the surface of LB solid medium, culture at 37°C for 16 hours, observe the colony formation and take pictures.
[0041] 2. Experimental results
[0042] image 3 It was shown that the growth state of the JM-dap strain containing the D...
Embodiment 3
[0045] Example 3 dap gene (DRB0118) expression in tobacco and salt tolerance identification of transgenic plants
[0046] (1) Construction of expression vector containing target gene
[0047] According to the full-length coding sequence, design primers to amplify the complete coding reading frame, and introduce restriction endonuclease sites on the forward and reverse primers respectively (this can be determined by the vector selected), so as to construct the expression vector. Using the amplified product obtained in Example 1 as a template, after PCR amplification, the sequence cDNA was cloned into an intermediate vector (such as pBluescript), and further cloned into a binary expression vector (such as pBI121 and pCAMBIA2200). The expression vector identified under the premise was transformed into Agrobacterium, and the model plant Nicotiana tabacum was transformed by the leaf disc method.
[0048] (2) Transforming Tobacco Using the Leaf Disk Method
[0049] Use a sterile t...
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