A method for detecting ploidy of watermelon

Abstract

The invention discloses a method for detecting the ploidy of watermelon. The steps include preparing protoplasts, extracting total DNA, and detecting DNA concentration by quantitative PCR. Protoplastsare isolated from leaves of watermelon with different ploidy. Total DNA is extracted from the same amount of protoplasts. According to the 5S rDNA sequence of watermelon, specific primers are designed for quantitative PCR. The standard curve of Ct values and DNA concentrations is plotted with different DNA concentrations. The concentration of Ct is converted and ploidy is calculated according toDNA concentration of different ploidy of watermelon. The method has low requirements for samples, and other plant tissues other than fruits and seeds with more nutrients can be separated from protoplasts. Fluorescence quantitative polymerase chain reaction (FQ-PCR) instrument with low requirement and low cost can meet the demand without purchasing relatively expensive Flow Cytometry (FCM). The accuracy of the method is similar to that of flow cytometry, and the precision of detection is higher than that of stomatal guard cell chloroplast counting method, and the operation flow is easier to realize than that of chromosome counting method. It provides a new way for watermelon ploidy detection.

Description

technical field [0001] The invention relates to the technical field of ploidy detection, in particular to a method for detecting ploidy of watermelon. technical background [0002] China is the largest country in watermelon production and consumption, and watermelon is one of the important economic crops in our country. Seedless watermelon is widely favored by consumers for its convenient eating characteristics, and occupies an important position in watermelon production. Seedless watermelon seed production is cumbersome and needs to be obtained by crossing diploid and tetraploid materials. Therefore, accurate and efficient ploidy detection methods are particularly important for seedless watermelon seed production and parent creation. At present, the commonly used cell ploidy detection method is flow cytometry detection method, which is easy to operate, but has high requirements for samples. The detection is prone to miscellaneous peaks, which will affect the detection res...

AI Technical Summary

Problems solved by technology

At present, the commonly used cell ploidy detection method is flow cytometry detection method, which is easy to operate, but has high requirements for samples. The detection is prone to miscellaneous peaks, which will affect the detection results

At the same time, the price of flow cytometer is high, and it is widely used in medical testing. The conventional plant research laboratory is less equipped, and the price of related reagent consumables is high, and the daily management and maintenance are more cumbersome.

In addition, the chromosome counting method is also one of the more accurate methods for ploidy identification, but it has high requirements for production, and a large number of chromosome production work is often required to obtain ideal chromosome sections

In contrast, molecular biology detection methods based on molecular markers have lower requirements for samples and equipment, such as GISH, FISH and other methods have been successfully applied to plant ploidy detection, but there is no search for quantitative PCR methods for ploidy detection. sex test reports

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