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Application of interfering lcyb and lcye expression and overexpressing egl3 in preparing Brassica plants with red petals

A technology of overexpression, Brassica, applied in the field of genetic engineering

Inactive Publication Date: 2017-06-23
SOUTHWEST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the entire Brassica genus including rapeseed, there is no report on the use of molecular breeding for metabolic engineering of rapeseed flower color.

Method used

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  • Application of interfering lcyb and lcye expression and overexpressing egl3 in preparing Brassica plants with red petals
  • Application of interfering lcyb and lcye expression and overexpressing egl3 in preparing Brassica plants with red petals
  • Application of interfering lcyb and lcye expression and overexpressing egl3 in preparing Brassica plants with red petals

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1. Microscopic observation of chromogenic subcellular organelles in Brassica and radish petals

[0042] Take fresh Brassica napus yellow petals (BnY); Brassica napus milk petals (BnW); Brassica napus yellow petals (BjY); Brassica napus yellow petals (BrY); White petals (BoW) and radish red petals (RsR) were stored on ice and shipped back to the laboratory, then sliced ​​by hand, observed with a low-power lens for quick screening, and qualified slices were selected for careful observation and photography. The results are as follows figure 1 shown. The results showed that the yellow substances in the yellow petals of several species of Brassica appeared as numerous small granules in one cell, distributed unevenly in the cell, and were squeezed to the wall-adhering position by the vacuoles, indicating that the yellow petals The chromogenic organelles are chromoplasts, whose pigments are yellow carotenoids. There are also some cells in the milky petals of Brassica...

Embodiment 2

[0043] Embodiment 2, detect brassica and radish petal pigment

[0044] In the morning of the full flowering period, the newly opened Brassica napus yellow petals (BnY); Brassica napus milk petals (BnW); Brassica napus yellow petals (BjY); BoY), kale white petals (BoW) and radish red petals (RsR), immediately transported back to the laboratory in a cool and fresh place, dried at 50-60°C, ground into powder, passed through a 60-mesh sieve, and stored in a dry place away from light.

[0045] 1. Test results of petroleum ether, hydrochloric acid and ammonia water

[0046] Weigh 0.100g of preserved petal powder, put them into numbered stoppered test tubes, add about 10mL of petroleum ether, 10% hydrochloric acid, and 30% ammonia water, mix gently, filter, and observe the color change. The results are as follows :

[0047] (1) Petroleum ether reaction: Brassica napus, Chinese cabbage rape, and mustard rape all show bright yellow, indicating that the content of carotenoids is high;...

Embodiment 3

[0067] Example 3. Molecular Identification of Brassica and Radish Petal Pigment Biosynthetic Pathways

[0068] (1) Expression characteristics of flavonoid pathway genes in Brassica and radish petals

[0069] In order to study the molecular mechanism of the difference in petal pigments between Brassica and radish, RT-PCR detection primers for the flavonoid pathway genes of Brassica and radish were designed, and 5SrRNA was used as an internal reference, as shown in Table 1:

[0070] Table 1. Primers for RT-PCR detection of flavonoid pathway

[0071]

[0072]

[0073]In the morning of the full flowering period, the newly opened Brassica napus yellow petals (BnY); Brassica napus milk petals (BnW); Brassica napus yellow petals (BjY); BoY), kale with white petals (BoW) and red radish petals (RsR), transported frozen in liquid nitrogen, and stored in a -80°C refrigerator for later use. Then use the RNAprep Pure plant total RNA extraction kit to extract the total RNA, after el...

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Abstract

The invention discloses an application of LCYB (lycopene-beta-cyclase) and LCYE (lycopene-epsilon-cyclase) expression interference and EGL3 excessive expression in preparation of brassica plants with red petals, and further discloses an expression vector for interfering LCYB and LCYE gene expression and simultaneously excessively expressing EGL3 genes and a preparation method of the expression vector. Yellow carotenoid is inhibited and red lycopene is accumulated through interference for LCYB and LCYE gene expression, accumulation of anthocyanin is promoted through excessive expression of EGL3 genes, so that petals are red, the brassica plants with the red petals are prepared successfully, and the flower colors of the brassica plants are enriched.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and specifically relates to the application of interfering with the expression of LCYB and LCYE while overexpressing the EGL3 gene in preparing brassica plants with red petals. Background technique [0002] Rapeseed is an important oil crop with a long history of cultivation, high economic value, wide application and strong adaptability. It is the largest oil crop in my country and an important oil crop worldwide. In terms of species taxonomy, rape is composed of three species of Brassica, Brassica napus (2n=38, AACC), Brassica rapassp.oleifera syn.B. campestris, 2n=20, AA), Brassica juncea (2n=36, AABB). Brassica napus is a compound species evolved from natural interspecific hybridization of Chinese cabbage (Brassica rapa) and cabbage (Brassica oleracea, 2n=18, CC), although the history of cultivation is only a few hundred years , but because of its strong growth potential and high yield, it ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/84A01H5/02
Inventor 柴友荣廖霏霏董博刘雪尹能文王君刘欢卢婧唐章林张梦琴
Owner SOUTHWEST UNIV
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