Method for detecting purity of GFP transgene cotton crossbreed

Abstract

The invention discloses a method for detecting the purity of a GFP transgene cotton crossbreed, which comprises the following steps: using a GFP homozygote variety as a male parent and a non-GFP variety as a female parent to obtain an F1 crossbreed as a detection object; extracting an F1 generation seed detection sample, soaking the seed in mild water of which the temperature is between 26 and 30DEG C for 20 to 30 hours, then accelerating germination at a temperature of between 28 and 32 DEG C for 20 to 30 hours, removing an external seed coat, and leaving and taking a hypocotyl and an internal seed coat of the same seed as a slice which is placed on the same glass slide respectively; irradiating the slices by blue-violet light of a fluorescence microscope, observing the fluorescence performances of the two slices, and determining that the seed is the F1 crossbreed when only the hypocotyl slice has green fluorescent light but the internal seed coat slice has no green fluorescent light; determining that the seed is a male parent or an F2 generation seed of GFP when both the hypocotyl slice and the internal seed coat slice have the green fluorescent light; and determining that the seed is a female parent or other varieties when both the hypocotyl slice and the internal seed coat slice have no green fluorescent light. The detection method is accurate, quick and economic, and is convenient for application.

Description

Technical field [0001] The invention belongs to the technical field of biological breeding, and specifically relates to a method for detecting the purity of GFP transgenic cotton hybrids. technical background [0002] Heterosis is a universal law in biology. The utilization of hybrid vigor is a hot spot in biological breeding. The wide application of the hybrid advantages of rice and corn has played a huge role in solving food security and feed safety. The utilization of cotton hybrid vigor started relatively late. At present, India and China are the countries with the largest application area of ​​cotton hybrids. The highest yield increase of cotton hybrids bred in India can reach more than 50%, and the promotion area accounts for about half of the cotton planting area. my country is a major exporter of traditional textiles. Cotton occupies an important position in my country's textile industry and agricultural production. The annual cotton planting area is about 60 million m...

AI Technical Summary

Problems solved by technology

The disadvantages of dominant morphological markers controlled by endogenous genes are: the range of parental selection is small, and it is difficult to breed high-dominant combinations.

Use kanamycin to identify the purity of cotton hybrids. The male parent of the hybrid must be Bt transgenic insect-resistant cotton, and the female parent must be non-Bt transgenic cotton. In the actual production of cotton hybrids, the female parent is the harvested plant, and the male and female The planting ratio is about 1:8. Because the female parent is not resistant to insects, the yield of seed production is greatly affected. Although the purity identification rate is above 90%, it still does not meet the purity standard of hybrid cotton seeds.

Kanamycin method can not detect other Bt transgenic cotton species mixed in F1

[0007] To sum up, the current cotton hybrid seed production mainly adopts manual hybrid seed production method. The loophole of this seed production method is: the plants with early flowering before emasculation have already formed selfing bolls, and the flower leakage and rainy days in the seed production process are also difficult. It is easy to form selfing bolls. The selfing bolls formed by late buds that have not been cleaned up after seed production are not completely peeled off and easy to form selfing seeds. The above conditions have a great impact on the purity of hybrids; in addition, the purchase price of cotton hybrids is very high. High, human confounding occurs from time to time

Cotton not only has low seed production efficiency and difficult to guarantee the purity of seed production, but also lacks accurate, fast and economical detection methods for the purity of hybrids. Therefore, the efficiency of seed production and detection methods for purity have become the technical bottleneck for the application of cotton hybrids

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