Genetic Engineering Application of Soybean Ribosomal Protein Coding Gene gmrpl12
A ribosomal protein and gene-encoding technology, applied in genetic engineering, application, plant genetic improvement, etc., can solve problems such as inability to predict functions, differences, etc.
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0023] 1) Cloning of soybean ribosomal protein coding gene GmRPL12
[0024] Take the soybean variety Kefeng No. 1 as the sample object, take its root, grind it with a mortar, add it to a 1.5mL EP tube filled with lysate, shake it fully, transfer it to a 1.5mL EP tube, and extract the total RNA (Total RNA Kit (Tiangen, Beijing, China). Use formaldehyde denaturing gel electrophoresis to identify the total RNA quality, and the spectrophotometer measures the RNA content. The total RNA obtained is a template, and the reverse transcription kit (TaKaRa Primer Script TM RT reagent kit, Japan) instructions for reverse transcription, after obtaining the first strand of cDNA, PCR amplification, the PCR program is as follows: 95 ° C pre-denaturation for 3 minutes, 95 ° C denaturation for 15 seconds, 60 ° C annealing for 15 seconds , 72°C extension for 45 seconds, a total of 35 cycles, and finally 72°C for 5 minutes, followed by a constant temperature of 12°C to obtain the cDNA of Kefeng...
Embodiment 2
[0035] 1) Construction of plant expression vectors
[0036] Design primers containing BamHI and KpnI double restriction site adapters and only the full length of the candidate gene, and use the CDS plasmid containing the target gene as a template for PCR amplification (see SEQ ID NO.9 and SEQ ID NO.10 for primer sequences) . The PCR program is as follows: pre-denaturation at 95°C for 3 minutes, denaturation at 95°C for 15 seconds, annealing at 60°C for 15 seconds, extension at 72°C for 45 seconds, a total of 35 cycles, and a final incubation at 72°C for 5 minutes, followed by constant temperature at 4°C. The CDS sequence of the soybean GmRPL12 gene with a length of 576 bp of the complete coding region is shown in SEQ ID NO.1 for its sequence. The amplified product and pMDC83 empty load were digested with BamHI and KpnⅠ, and the digested vector was purified. Combine the PCR product with the pMDC83 linear vector after enzyme digestion according to Novizyme Entry One Step Clo...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More