CN-121204280-B - Primer and method for identifying whether citrus fruits have low-temperature color-promoting potential

CN121204280BCN 121204280 BCN121204280 BCN 121204280BCN-121204280-B

Abstract

The invention discloses a primer and a method for identifying whether citrus fruits have low-temperature color-promoting potential. After PCR amplification reaction is carried out on DNA of citrus plants by designing specific primers aiming at CitHB promoter, whether fruits have postharvest low-temperature treatment color promotion potential is judged according to amplified target bands. Citrus plants with 750 bp target bands, the fruits of which have postharvest low temperature color development potential, citrus plants with 100 bp target bands, the fruits of which do not have low temperature color development potential. The method provided by the invention can be used for rapidly identifying whether the citrus fruits have low-temperature color-promoting potential, and provides a guarantee for the accurate application of the low-temperature color-promoting technology in the postharvest treatment of the citrus fruits.

Inventors

DENG XIUXIN
YE JUNLI
WEI RANRAN
Zhong Guoxiu
ZHU SHENCHAO
TANG JIN
FANG YU

Assignees

华中农业大学

Dates

Publication Date: 20260512
Application Date: 20251010

Claims (6)

1. A primer for identifying whether citrus fruits have postharvest low-temperature color-promoting potential is characterized by comprising a primer combination of a nucleotide sequence 1F and a sequence 1R, wherein the nucleotide sequence 1F is shown as SEQ ID.1, and the nucleotide sequence 1R is shown as SEQ ID.2.
2. A detection reagent, kit or chip comprising the primer of claim 1.
3. Use of the detection reagent, kit or chip of claim 2 for identifying whether citrus fruit has post-harvest low temperature color development potential.
4. A method for identifying whether citrus fruit has post-harvest low temperature color development potential comprising the steps of: step1, extracting genome DNA of citrus plants to be detected; step2, performing PCR amplification by using the genomic DNA extracted in the step 1 as a template and using the primer, wherein the primer is the primer in claim 1; step 3, parting the PCR amplification product obtained in the step2, and carrying out strip discrimination on the parting result: If the product with the size of 750 bp is obtained through amplification, the citrus variety is judged to have the low-temperature color-promoting potential after the citrus variety is picked, the color of the peel can be improved by utilizing the temperature-treated color-promoting technology after the citrus variety is picked, and if the product with the size of 100 bp is obtained through amplification, the citrus variety is judged to not have the low-temperature color-promoting potential, and the low-temperature color-promoting technology is not recommended to improve the color of the peel.
5. The method according to claim 4, wherein the PCR amplification reaction system in step 2 comprises 10. Mu.L of 2X TAQ MASTER Mix, 0.5. Mu.L of nucleotide sequence 1F, 0.5. Mu.L of nucleotide sequence 1R, 1.0. Mu.L of genomic DNA of the citrus plant to be tested and 8. Mu.L of ddH 2 O, and the concentrations of the solutions of the nucleotide sequence 1F and the nucleotide sequence 1R are 10. Mu.mol/L.
6. A method for identifying citrus fruit having low temperature color development potential according to claim 5, wherein the PCR amplification is performed under conditions of 95℃initial pre-denaturation of 5min followed by 34 cycles each comprising 95℃denaturation of 30 sec, 58℃annealing of 30 sec and 72℃extension of 30 sec, 72℃final extension of 5min after the end of the cycle and final 4℃preservation.

Description

Primer and method for identifying whether citrus fruits have low-temperature color-promoting potential Technical Field The invention belongs to the technical field of plant genetic engineering, and particularly relates to a primer and a method for identifying whether citrus fruits have postharvest low-temperature color-promoting potential. Background Citrus is one of the most important economic fruit trees worldwide, and the appearance and color of fruits are key indexes for evaluating commodity value. Carotenoids are the primary pigments that make up the color of citrus peel. In production practice, the harvested citrus fruits are subjected to proper low-temperature treatment, so that synthesis and accumulation of carotenoid can be effectively promoted, the color of peel is remarkably improved, and the market competitiveness is improved. However, post-harvest low temperature color-promoting techniques are not effective for all citrus varieties. At present, whether a variety has low-temperature color promotion potential is judged, and the traditional postharvest physiology method is completely relied on, namely, the actual low-temperature treatment test is carried out after fruits are picked, so that the color change is observed. The whole verification process of the method takes weeks to months, can not provide basis for post-harvest treatment decision in time, and greatly limits the accurate application of the low-temperature color-promoting technology. In recent years, research is being carried out in an attempt to explore the physiological and molecular mechanisms of low-temperature color development, but no key molecular marker capable of being directly and effectively used for identifying the postharvest low-temperature color development capability of citrus varieties has been discovered. Therefore, the technical scheme capable of rapidly and accurately identifying whether the variety has low-temperature color-promoting potential before or during fruit harvesting is urgently needed in the field so as to overcome the defect that the prior art needs to rely on post-harvest physical verification and realize the on-demand and efficient application of post-harvest treatment technology. Disclosure of Invention In order to solve the technical problems, the invention mainly aims to provide the primer and the method for identifying whether the citrus fruits have the low-temperature color-promoting potential after being picked so as to quickly, accurately and simply identify whether the citrus fruits have the low-temperature color-promoting potential and provide an efficient auxiliary tool for quality management after being picked. In order to achieve the purpose of the invention, the invention provides the following technical scheme: In a first aspect, a primer for identifying whether citrus fruits have postharvest low-temperature color-promoting potential is a primer combination comprising a nucleotide sequence 1F and a sequence 1R, wherein the nucleotide sequence 1F is shown in SEQ ID.1, and the nucleotide sequence 1R is shown in SEQ ID.2. In a second aspect, the invention also provides a detection reagent, kit or chip containing the primer. Further, the detection reagent, kit or chip also comprises other reagents for PCR amplification. In a third aspect, the detection reagent, kit or chip of the invention can be used to identify whether citrus fruits have low temperature color development potential. In a fourth aspect, the present invention also provides a method for identifying whether citrus fruit has post-harvest low temperature color development potential, comprising the steps of: Step1, extracting genome DNA of citrus plants to be detected, Step 2, performing PCR amplification by using the genomic DNA extracted in the step 1 as a template and using a primer, wherein the primer is a primer combination comprising a nucleotide sequence 1F and a sequence 1R, the nucleotide sequence 1F is shown as SEQ ID.1, and the nucleotide sequence 1R is shown as SEQ ID.2; step 3, parting the PCR amplification product obtained in the step2, and carrying out strip discrimination on the parting result: if the product with the size of 750 bp is obtained through amplification, the citrus variety is judged to have the low-temperature color-promoting potential after picking, the color of the peel can be improved by utilizing the low-temperature color-promoting technology, and if the product with the size of 100 bp is obtained through amplification, the citrus variety is judged to not have the low-temperature color-promoting potential, and the low-temperature color-promoting technology is not recommended to improve the color of the peel. Further, the PCR amplification reaction system of step 2 comprises 10. Mu.L of 2X TAQ MASTER Mix, 0.5. Mu.L of nucleotide sequence 1F, 0.5. Mu.L of nucleotide sequence 1R, 1.0. Mu.L of genomic DNA of the citrus plant to be tested and 8. Mu.L of ddH, and the solution concentrations of the nucleo