A kind of peony plmyb108 gene and its application in plant drought resistance
A peony gene technology, applied in the field of peony PlMYB108 gene and its application in plant drought resistance, to achieve the effect of enhancing drought resistance
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Embodiment 1
[0046] Cloning of Example 1 Peony PlMYB108 gene cDNA full-length sequence
[0047] Acquisition of the 3′-terminal cDNA sequence of PlMYB108 gene: 5-year-old peony 'Dafugui' potted seedlings (loam: peat: coarse sand = 1:1:1) were used as materials for natural drought stress treatment, and the 21-day-old peony was treated with drought stress Leaves were used as materials, and total RNA was extracted using the MiniBEST Plant RNA Extraction Kit (TaKaRa) kit. Use 3′full RACECore Set Ver.2.0 (TaKaRa) to reverse transcribe the first strand of cDNA. The reverse transcription system is: 1μL RNA, 1μL 3′-RACE Adapter, 1μL dNTP Mixture (10mM each), 2μL 5× M-MLV Buffer, 0.25 μL RNase Inhibitor, 0.25 μL Reverse Transcriptase M-MLV (RNase H - ), 4.5 μL RNase Free ddH 2 O; Reverse transcription program: react at 42°C for 60 minutes, and extend at 70°C for 15 minutes. On this basis, 3′-RACE amplification was completed by two rounds of PCR. The first round of PCR amplification system is: 2 ...
Embodiment 2
[0049] The expression of embodiment 2 Paeoniae officinale overexpression vector pBWA(V)HS-PlMYB108-GLosgfp in tobacco
[0050] Construction of peony PlMYB108 overexpression vector pBWA(V)HS-PlMYB108-GLosgfp: The obtained full-length sequence of PlMYB108 gene was sent to Wuhan Boyuan Biotechnology Co., Ltd. for whole gene synthesis, and cDNA template was obtained, and then PCR amplification was carried out. For: 4 μL Mg 2+ , 2 μL dNTP Mixture (2.5mM each), 1 μL cDNA template, 2 μL PlMYB108-Forward Primer (5′-CAGTGGTCTCACAACATGGATGTTAATGGGAGAGG-3′(SEQ ID NO.4)), 2 μL PlMYB108-Reverse Primer (5′-CAGTGGTCTCATACAAATATTSETGCTGGAGAACTGTT-3 NO.5)), 5μL 10×LA PCR Buffer II, 0.5μL TaKaRa LA Taq (5U / uL) and 33.5μL ddH 2 O. The amplification program was: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 30 s, annealing at 50°C for 45 s, extension at 72°C for 52 s, and 30 cycles; extension at 72°C for 10 min, and 16°C for 30 min. Cut out the target fragment, perform gel recov...
Embodiment 3
[0057] Determination of Relative Water Content of Leaf of Tobacco Plant under Embodiment 3 Drought Stress
[0058] The weight of an appropriate amount of fresh leaves weighed by a balance (BSA224S, Sartorius, Germany) was recorded as fresh weight (FW), and treated in an oven at 105°C for 5 minutes, and then adjusted to 65°C for more than 2 hours, and then weighed and dried The sample with constant weight is recorded as dry weight (DW), and the relative water content of leaves is calculated according to the following formula: relative water content of leaves (%)=(FW-DW) / FW×100%. From Figure 5 It can be seen that compared with the wild-type tobacco, the transgenic tobacco of PlMYB108 has significantly higher leaf relative water content.
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