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Method for regulating cloning and transcription of lignin metabolism-related ethylene response factors (ERFs) in Eriobotrya japonica Lindl.
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A response factor, lignin technology, applied in the field of loquat lignin metabolism-related ethylene response factor cloning and transcriptional regulation, can solve problems such as unclear biological mechanism
Active Publication Date: 2015-07-15
ZHEJIANG UNIV
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Problems solved by technology
The biological mechanism of ERFs family members regulating lignification during low-temperature storage of loquat fruit is still unclear
Method used
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Embodiment 1
[0021] Example 1 Expression of loquat ERF in programmed cooling (LTC)
[0022] (1) Experimental method
[0023] 1. Material treatment: Select Luoyangqing (LYQ), a red-fleshed loquat fruit with uniform size, no pests and injuries, and no scratches. One batch is directly stored at 0°C, and the other batch is subjected to programmed cooling (LTC), that is, at 5°C. After 6 days of storage, transfer to 0°C for storage.
[0024] 2. The firmness of loquat fruit was measured with a texture analyzer (Stable Micro Systems, UK). Each fruit was measured at the equator of the symmetrical plane of the fruit, and the average value was taken as a repetition, and a single fruit was repeated 10 times. The coefficient of determination is as follows: probe P5, penetration depth 4 mm, pre-test speed 3 mm s -1 , with a measurement speed of 1 mm s -1 , the measured speed is 10 mm s -1 . The final data is expressed in Newtons (N).
[0025] 3. Determination of lignin content. Washing of samples...
Embodiment 2
[0030] Example 2 Expression of relevant ERFs in heat shock (HT)
[0031] (1) Experimental method
[0032] (1) Material treatment: choose Luoyangqing (LYQ), a red-fleshed loquat fruit with the same size, no pests and abrasions, and store one batch directly at 0°C, while the other batch is subjected to hot air treatment at 40°C (heat shock, HT ) After 4 hours, transfer to 0°C for storage.
[0033] (2) The firmness of loquat fruit was determined by a texture analyzer (Stable Micro Systems, UK), and the lignin content was determined (as above).
[0034] (3) The fruit RNA was extracted by the CTAB method, and then DNAcontamination was removed, and reverse transcription was performed. Quantitative PCR analysis was performed on the obtained 32 ERF Unigenes.
[0035] (2) Experimental results
[0036] (1) Compared with the fruit directly stored at 0°C, the increase of fruit firmness and lignin content under HT treatment was slowed down, see attached figure 1 (A), (B).
[0037] ...
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Abstract
The invention provides a method for regulating the cloning and the transcription of ligninmetabolism-related ERFs in Eriobotryajaponica Lindl.. The method for regulating the transcription of the ERFs in the ligninanabolism is carried out through obtaining an ERF sequence fragment from deep Eriobotryajaponica Lindl. sequencing results and obtaining the 3' terminal sequences of the ERFs. According to the invention, the deep sequencing results are cloned to the 3' terminal nucleotide sequence of an ERF Unigene in the Eriobotryajaponica Lindl. to obtain thirty-two 3' terminal sequences, and the changes of ERF expression levels of Eriobotrya japonica Lindl. fruits of the ERFs processed at 0DEG C, HT and LTC are obtained to obtain two ERF gene family members related with cold injury lignification, so basal theoretical bases are provided for the further researches of the ERF action mechanism and fresh keeping technologies of picked fruits.
Description
technical field [0001] The invention belongs to the technical fields of fruit storage and plantmolecular biology, and relates to a method for cloning and transcription regulation of loquat ligninmetabolism-related ethylene response factors. Background technique [0002] Loquat ( Eriobotrya japonica Lindl. ) is an evergreenfruit tree of the Rosaceae native to my country, which belongs to non-climtic fruit. Different from the postharvestsoftening process of other fruits, red meat loquats will undergo lignification during postharvest storage, resulting in increased fruit firmness, weak flavor, rough and less juice, etc., seriously affecting fruit quality and economic value. The low temperature of 0-5℃ can significantly inhibit the respiration and ethylene release of loquat fruit, thus prolonging the shelf life, but the chilling injury of the fruit will occur at 0-1℃, which will accelerate the lignification of loquat fruit. Programmed cooling (LTC), that is, storing loquat...
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