Application of wheat flavone-5-O-glucoside in regulating and controlling resistance of plants to weeds
A technology of ticlavone and glucose, applied in the field of application of ticlavone-5-O-glucoside in regulating plant resistance to weeds, can solve problems such as genetic mechanism and weed suppression mechanism are not very clear
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Embodiment 1
[0055] The compound was purified Example 1,
[0056] Rice (Indica) after about 60 days (1) collecting the fresh leaves sowing, triturated after freeze drier powder. Adding 8 volumes of 75% ethanol, ultrasonic extraction three times, each time 20min.
[0057] (2) The extract was filtered and concentrated to alcohol-free flavored by a rotary evaporator, followed by addition of an equal volume of petroleum ether, methylene chloride, and extracted three times with n-butanol respectively.
[0058] (3) The obtained n-butanol (Biotage Isolera concentrated using a rotary evaporator in a low pressure flash with a phase preparative liquid chromatography TM Prime) on the sample using a wet coarse fraction. Silica gel column YMC * GEL C18 packing of spherical filler, the filler particle diameter of 50 m, a pore size of 12nm. In this embodiment the filler weight of 100g. Water mobile phase (containing 0.1% formic acid, expressed in% volume percentage), methanol. The mobile phase was methanol r...
Embodiment 2
[0065] Example 2, Os07g0503900 Clone
[0066] (1) (Oryza sativa L.ssp.japonica cv.Zhonghua11, ZH11) Genomic DNA extraction of the leaves of rice seedlings in flower 11.
[0067] (2) According to the existing sequence information Os07g0503900 site annotation rice (Rice Genome Annotation Project), with primers containing BamHI and HindШ of F1 / R1 to ZH11 was amplified genomic DNA sequences containing CDS Os07g0503900 gene enzyme sites.
[0068] Primer F1: 5'-ggatccATGGCTCCAGCGATGGCGAG-3 ';
[0069] Primer R1: 5'-aagcttCTATATGGATGACATGTGGGC-3 '.
[0070] (2) The gene fragment was ligated to pEASY-T 3 Carrier (TransGen Biotech, -T3Cloning Kit), to transform E. coli DH5α competent cells, using blue-white screening positive clones.
[0071] (3) The positive clones using primers F2 / R2 identified PCR, the amplified fragment was 310bp for the positive clones. The primers are as follows:
[0072] Primer F2: 5'-AGGACTTCATCTCCCGGTTCATGC-3 ';
[0073] Primer R2: 5'-TTCTTCATCACAGGCGTCGGCAAC...
Embodiment 3
[0076] Application Example 3, Os07g0503900 gene embodiments
[0077] A prokaryotic expression vector
[0078] (1) cut with the restriction enzymes BamHI and HindШ recombinant vector of Example 2 obtained pEASY-T 3 -Os07g0503900 complete digestion, while cut expression vector pMAL-c2X (Chinese overseas, VECT-570). System for the digestion: 5μg plasmid, 2.5μL 10 × digestion buffer, 2μL BamHI, 2μL HindШ, add ddH 2 O complement reaction system to 50μL. Digestion reaction conditions are: enzyme 37 ℃ 4 hours.
[0079] (2) digested products by agarose gel electrophoresis to separate and recover a size of about 1.5Kb fragment containing Os07g0503900, and approximately 6.6Kb pMAL-c2X vector fragment were dissolved in 30μL ddH 2 O.
[0080] (3) The step (2) were obtained gene fragment with a vector backbone fragment ligation reaction. Ligation reaction is: 1μL 10 × ligase buffer, 0.5μL T 4DNA ligase, 1μL PMAL-C2X vector fragment, 3 μl of gene fragment, DDH 2 O Supplement the reaction system...
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