Primers, kits and applications for controlling wheat leaf senescence
A technology of leaf senescence and wheat, which is applied in the fields of plant genetic engineering and wheat molecular breeding, can solve the problems of being difficult to be used and poor agronomic traits
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[0055] 1. TaNACS gene cloning and overexpression vector construction
[0056] Using Chinese spring wheat cDNA as template, PCR amplification was carried out with SEQ ID NO.1 as upstream primer and SEQ ID NO.2 as downstream primer. The template concentration was 50ng / ml, and the amplification procedure was as follows:
[0057] 95°C for 3min; 33 cycles of 95°C for 30s, 59°C for 30s, 72°C for 1min; 72°C for 10min.
[0058] The PCR amplified product was connected to the pCambia3301 overexpression vector (ubi-driven overexpression) by infusion recombinase, finally forming the recombinant vector pCambia3301-TaNACS.
[0059] The connection method is:
[0060] (1) Digest the pCambia3301 vector with EcoRI and BamHI, and recover the fragments for later use;
[0061] (2) Establish the ligation system: 1.5 μL of the digested pCambia3301 vector; 2.5 μL of PCR fragments; 1 μL of infusion enzyme:
[0062] (3) Establish reaction conditions: 50°C for 15 minutes, place on ice for 5 minutes; ...
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