An Effective Method for Detecting Genes Induced by Bacterial Blight in Rice
A technique for inducing expression, bacterial blight, applied in the field of molecular genetics
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Embodiment 1
[0026] 1. Preparation method of inoculum:
[0027] The bacterial blight PXO86 was inoculated on the PDA medium (potato 300g / L, glucose 30g / L, agar 15g / L), at 28 Cultivate in a warm incubator for 3-4 days, wash the colonies from the culture medium with distilled water, and dilute to a concentration of 9×10 8 cfu / mL of bacterial suspension. During the tillering stage of rice, the leaves of the resistant rice variety IRBB7 and the susceptible rice variety IR24 were infected with bacterial blight by the leaf cutting method (Kauffman et al., 1973). After infecting for 48 hours, select I to V area ( figure 1 ) leaf tissues were quickly frozen in liquid nitrogen at -80 live.
[0028] instruction manual:
[0029] Area I is 0 to 1 cm from the notch, area II is <1 to 2 cm from the notch, area III is <2 to 3 cm from the notch, area IV is <3 to 5 cm from the notch, and area IV is <3 to 5 cm from the notch. <5 to 7cm is area V.
[0030] Remarks: When sampling each of the above are...
Embodiment 2
[0045] Example 2. Change the gene (Os03g0853200) in Example 1 to OsPR1b (GenBankaccession: EF061247.1), change the corresponding primer sequence to 5'-GGCAACTTCGTCGGACAGA-3' and 5'-CCGTGGACCTGTTTACATTTTCA-3', and the rest are the same Example 1.
[0046] Remarks: After verification by conventional methods, the obtained gene is indeed OsPR1b.
[0047] Experimental results such as image 3 As shown, in the susceptible variety IR24, the gene was not induced by bacterial blight; while in the I region of the inoculated resistant variety IRBB7 leaves (that is, within 1cm from the cut), the gene was most significantly detected The gene is induced to express.
[0048] Remarks: The rice disease resistance-related gene OsPR1b induced by pathogenic bacteria is a recognized disease resistance marker gene, which is used to indicate the activation of plant disease resistance system.
Embodiment 3
[0049] Example 3. Change the gene (Os03g0853200) in Example 1 to OsNPR1 (GenBankaccession: DQ450948.1), change the corresponding primer sequence to 5'-CTGATCCGGTTTCCCTCGGA-3' and 5'-GACCTGTCATTCTCCTCCTTG-3', and the rest are the same Example 1.
[0050] Remarks: After verification by conventional methods, the obtained gene is indeed OsNPR1.
[0051] Experimental results such as Figure 4 As shown, in the susceptible variety IR24, the gene was not induced by bacterial blight; while in the I region of the inoculated resistant variety IRBB7 leaves (that is, within 1cm from the cut), the gene was most significantly detected The gene is induced to express.
[0052] Remarks: The rice disease resistance-related gene OsNPR1 induced by pathogenic bacteria is also a recognized disease resistance marker gene, which is used to indicate the activation of plant disease resistance system.
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