Molecular marker of high quercetin 7-O-β-D-glucuronic acid content associated with root-knot nematode resistance and its application

Abstract

The present invention provides a molecular marker of high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease and its application, belonging to the field of molecular biotechnology, the SV molecular marker is Chr14_2657774_del_1565, which is used to predict whether the quercetin 7-O-β-D-glucuronic acid content in cassava is high or low; the primer set for detecting the SV molecular marker includes: an upstream primer sequence of 5'-ATGATCCGAGATCCAATG-3'; a downstream primer sequence of 5'-GTTACCCAAATGTGATGC-3'; a kit is prepared using the primer set; cassava is detected using the primer set or the kit to predict whether the quercetin 7-O-β-D-glucuronic acid content in cassava is high or low. The SV molecular marker of the present invention can accurately and efficiently predict whether the quercetin 7-O-β-D-glucuronic acid content in cassava is high or low during the cassava seedling stage, thereby improving breeding efficiency.

Description

Technical Field

[0001] The present invention belongs to the technical field of plant breeding, and relates to a breeding molecular marker method for the content of metabolites related to resistance to root-knot nematode disease, and a breeding technology for screening high-quercetin materials using an SV molecular marker related to the high quercetin 7-O-β-D-glucuronic acid (Quercetin 7-O-β-D-Glucuronide) content in cassava as a key indicator. The present invention specifically relates to a molecular marker for high quercetin 7-O-β-D-glucuronic acid content related to resistance to root-knot nematode disease and its application, which solves the problems of long breeding time and high breeding cost for new cassava varieties with high quercetin 7-O-β-D-glucuronic acid content. Background Art

[0002] Root-knot nematode disease is a disease caused by root-knot nematodes that primarily harm plant roots, severely impacting growth and yield. Root-knot nematodes primarily harm the root system, causing the formation of nodular root knots of varying sizes on lateral and fibrous roots. Plants infested by root-knot nematodes experience weak growth of the aboveground parts, dwarfed stature, and yellowing and curling leaves. In severe cases, leaves dry up and fall off, leading to a decrease in yield. Flavonoids may play an important role in a plant's anti-nematode response. Prevention and control of root-knot nematode disease is difficult and costly, and traditional prevention and control methods fail to fundamentally address the problem.

[0003] Flavonoids play an important role in plant resistance to nematodes. Quercetin, a natural flavonoid, exhibits multiple biological activities, including antioxidant, anti-inflammatory, and antimicrobial properties. Studies have shown that quercetin and its derivatives have been shown to inhibit root-knot nematodes. Quercetin 7-O-β-D-glucuronide, a metabolite of quercetin, is a metabolite of quercetin. Plants high in quercetin 7-O-β-D-glucuronide typically exhibit enhanced antioxidant activity and resistance to root-knot nematode stress, making it suitable for cultivation in areas with high root-knot nematode prevalence to enhance crop resistance.

[0004] In traditional cassava breeding, breeders primarily select individual plants with superior traits, fix these traits through hybridization or backcrossing, and then propagate superior seedlings through asexual reproduction. Currently, to obtain new cassava varieties with high quercetin 7-O-β-D-glucuronic acid content, it is necessary to wait for mature plants and then investigate the quercetin 7-O-β-D-glucuronic acid content in their roots to screen for superior plants. Due to the lack of molecular markers significantly associated with high quercetin 7-O-β-D-glucuronic acid content, this breeding method is time-consuming, costly, and inefficient. Summary of the Invention

[0005] In response to the above problems, the present invention provides a molecular marker of high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease and its application.

[0006] To achieve the above object, the technical solution adopted by the present invention is:

[0007] A molecular marker with high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease, the molecular marker being an SV molecular marker, the SV molecular marker being a large fragment of bases 2657774 to 2659338 of chromosome 14 of the cassava genome, with a sequence length of 1565 bp, named Chr14_2657774_del_1565, and the sequence being shown in SEQ ID NO: 1;

[0008] The quercetin 7-O-β-D-glucuronic acid content in cassava carrying the SV molecular marker was significantly higher (i.e., greater) than the quercetin 7-O-β-D-glucuronic acid content in cassava lacking the SV molecular marker (i.e., a large deletion at bases 2657774 to 2659338 on chromosome 14);

[0009] That is, the cassava carrying the SV molecular marker has a higher quercetin 7-O-β-D-glucuronic acid content, and is a cassava with a high quercetin 7-O-β-D-glucuronic acid content;

[0010] The cassava lacking this SV molecular marker has a lower quercetin 7-O-β-D-glucuronic acid content, and is low-quercetin 7-O-β-D-glucuronic acid content cassava.

[0011] The invention discloses an application of a reagent for detecting the molecular marker of high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease. The application is to detect whether cassava carries the SV molecular marker by using the reagent to predict whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low.

[0012] A primer set for detecting the above-mentioned high quercetin 7-O-β-D-glucuronic acid content molecular marker associated with root-knot nematode resistance, wherein the primer set is designed using the above-mentioned SV molecular marker, that is, the primers are designed based on nucleotides 2657774 to 2659338 of chromosome 14 of the cassava genome, and include:

[0013] The upstream primer sequence was 5′-ATGATCCGAGATCCAATG-3′;

[0014] The downstream primer sequence was 5′-GTTACCCAAATGTGATGC-3′.

[0015] A kit comprising the above primer set.

[0016] Furthermore, the kit includes reagents for PCR amplification.

[0017] Furthermore, the kit includes: 2× Rapid Taq Master Mix and ddH2O.

[0018] A method for identifying whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low, the method comprising: using the above-mentioned primer set and cassava genomic DNA as a template to perform PCR amplification on the SV molecular marker, sequencing the PCR amplification product, and determining whether the SV molecular marker is carried to predict whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low;

[0019] Alternatively, the SV molecular marker is amplified by PCR using the above kit and cassava genomic DNA as a template, and the PCR amplification product is sequenced to determine whether it carries the SV molecular marker, so as to predict whether the cassava has a high or low quercetin 7-O-β-D-glucuronic acid content;

[0020] When the nucleotide sequence of the PCR amplification product is as shown in SEQ ID NO: 1, and is 1565 bp in length, and the sequence is a DNA fragment represented by the SV molecular marker, that is, a sequence within the range of coordinates 1 to 1565 from the 5' end of the SV molecular marker, then the cassava carries the SV molecular marker, and the cassava has a high quercetin 7-O-β-D-glucuronic acid content, and is a cassava with a high quercetin 7-O-β-D-glucuronic acid content;

[0021] Otherwise, the cassava lacks the SV molecular marker, that is, a large fragment deletion occurs at bases 2657774 to 2659338 of chromosome 14 in the sample genome, and no amplification product is obtained by PCR amplification. In this case, the cassava has a low quercetin 7-O-β-D-glucuronic acid content, and is a cassava with low quercetin 7-O-β-D-glucuronic acid content.

[0022] Furthermore, the PCR amplification system includes: 12.5 μL of 2× Rapid Taq Master Mix, 1 μL of the upstream primer with a concentration of 10 μM, 1 μL of the downstream primer with a concentration of 10 μM, 1 μL of cassava genomic DNA, and 9.5 μL of ddH2O;

[0023] The reaction program of PCR amplification was as follows: 95°C for 5 min; 95°C for 30 s, 60°C for 30 s, and 72°C for 30 s, 35 cycles; and 72°C for 5 min.

[0024] A method for identifying whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low, the method comprising using the above-mentioned primer set and cassava genomic DNA as a template to perform PCR amplification on the SV molecular marker, performing electrophoresis detection on the PCR amplification product, and interpreting the electrophoresis detection result to determine whether the cassava carries the SV molecular marker, so as to predict the cassava quercetin 7-O-β-D-glucuronic acid content;

[0025] Alternatively, the SV molecular marker is amplified by PCR using the above kit and cassava genomic DNA as a template, the PCR amplification product is detected by electrophoresis, and the electrophoresis detection result is interpreted to determine whether the SV molecular marker is carried, so as to predict whether the cassava has a high or low quercetin 7-O-β-D-glucuronic acid content;

[0026] When only a 1565 bp band appears in the electrophoresis detection result, the cassava carries the SV molecular marker, and the cassava has a high quercetin 7-O-β-D-glucuronic acid content, which is a cassava with a high quercetin 7-O-β-D-glucuronic acid content;

[0027] Otherwise, when the electrophoresis detection result shows no band, the cassava lacks the SV molecular marker, and the cassava has a low quercetin 7-O-β-D-glucuronic acid content, which is low quercetin 7-O-β-D-glucuronic acid content cassava.

[0028] Furthermore, the PCR amplification system includes: 12.5 μL of 2× Rapid Taq Master Mix, 1 μL of the upstream primer with a concentration of 10 μM, 1 μL of the downstream primer with a concentration of 10 μM, 1 μL of cassava genomic DNA, and 9.5 μL of ddH2O;

[0029] The reaction program of PCR amplification was as follows: 95°C for 5 min; 95°C for 30 s, 60°C for 30 s, and 72°C for 30 s, 35 cycles; and 72°C for 5 min.

[0030] The beneficial effects of the molecular marker of high quercetin 7-O-β-D-glucuronic acid content related to resistance to root-knot nematode disease and its application are as follows:

[0031] The present invention discloses a molecular marker for high quercetin 7-O-β-D-glucuronic acid content associated with root-knot nematode resistance, solving the problems of long breeding time, high breeding costs, and low efficiency in the prior art for new cassava varieties related to quercetin 7-O-β-D-glucuronic acid content. The SV molecular marker occurs at base 2657774 of chromosome 14 (Chr14) of the cassava genome, with a sequence deletion of 1565 bp, and is named Chr14_2657774_del_1565. The SV molecular marker can accurately and efficiently predict high or low quercetin 7-O-β-D-glucuronic acid content in cassava fibrous and tuberous root tissues, greatly improving the efficiency of selecting root-knot nematode-resistant cassava varieties with high quercetin 7-O-β-D-glucuronic acid content and reducing costs.

[0032] This SV molecular marker can be used to accurately and efficiently predict the high or low quercetin 7-O-β-D-glucuronic acid content in cassava fiber roots and tubers during the cassava seedling stage by determining whether leaf DNA carries the target molecular marker. This allows for rapid screening of high-quality plants with high quercetin 7-O-β-D-glucuronic acid content during the cassava seedling stage, greatly improving the selection efficiency of cassava breeding and effectively assisting in cassava breeding selection, with extremely high economic value.

[0033] The present invention develops a molecular marker for high quercetin 7-O-β-D-glucuronic acid content associated with root-knot nematode resistance, and establishes a method for accurately and rapidly identifying whether cassava has a high or low quercetin 7-O-β-D-glucuronic acid content, providing a more effective theoretical and practical basis for molecular marker-assisted breeding of new cassava varieties with specific quercetin 7-O-β-D-glucuronic acid content levels. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is a Manhattan plot of the GWAS signal of the genome-wide chromosome association analysis positioning of the trait data of the relative content of quercetin 7-O-β-D-glucuronic acid in 337 samples of cassava in Example 1 of the present invention;

[0035] Figure 2 These are the T-test results for the relative content levels of quercetin 7-O-β-D-glucuronic acid in 207 samples of cassava in Example 5 of the present invention; wherein, REF represents cassava carrying the SV molecular marker, and n=113 represents a sample size of 113; SV represents cassava lacking the SV molecular marker, and n=94 represents a sample size of 94. DETAILED DESCRIPTION

[0036] The technical solutions in the embodiments of the present invention are described clearly and completely below. Many specific details are set forth in the following description to facilitate a full understanding of the present invention. However, the present invention may also be implemented in other ways different from those described herein, and those skilled in the art may make similar generalizations without departing from the scope of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below. The present invention is further described in detail below in conjunction with specific embodiments to facilitate understanding by those skilled in the art.

[0037] Example 1 Obtaining a molecular marker for high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease

[0038] We collected 337 fresh cassava leaf samples grown in Danzhou, Hainan Province, China (109.5°E, 19.5°N), extracted genomic DNA, and performed whole-genome sequencing. The high-quality sequencing data after quality control were aligned to the cassava reference genome. A genome-wide association study (GWAS) was performed using the resulting cassava genome-wide SV molecular marker map and phenotypic data on the relative content of quercetin 7-O-β-D-glucuronic acid in cassava germplasm resources. The results are as follows: Figure 1 As shown, it can be seen that there is a significant correlation signal with the content level of cassava quercetin 7-O-β-D-glucuronic acid on the cassava genome (indicated by the arrow), including one SV molecular marker (Chr14_2657774_del_1565).

[0039] A molecular marker for high quercetin 7-O-β-D-glucuronic acid content, associated with resistance to root-knot nematode disease, is located on chromosome 14 of the cassava genome. The 1565-bp long fragment, designated Chr14_2657774_del_1565, is shown in SEQ ID NO: 1. Testing of cassava breeding materials using this SV molecular marker revealed significantly higher quercetin 7-O-β-D-glucuronic acid content in cassava varieties carrying the SV molecular marker compared to those lacking the SV molecular marker (i.e., a large deletion of bases 2657774 to 2659338 on chromosome 14).

[0040] Therefore, the SV molecular marker can be used to accurately and efficiently predict the high or low content of quercetin 7-O-β-D-glucuronic acid in cassava fiber roots and tubers during the cassava seedling stage by checking whether the leaf DNA carries the target molecular marker. Excellent plants with high content can be quickly screened out during the cassava seedling stage, greatly improving the selection efficiency of cassava breeding, effectively assisting cassava breeding selection, and having extremely high economic value.

[0041] Example 2 Primer Set and Kit for Identifying High or Low Content of Quercetin 7-O-β-D-Glucuronic Acid

[0042] Using the characteristics of the high quercetin 7-O-β-D-glucuronic acid content molecular marker associated with root-knot nematode resistance in Example 1 on the cassava genome, a set of primers was designed (i.e., the primers were designed based on nucleotides 2657774 to 2659338 of chromosome 14 of the cassava genome), specifically as follows:

[0043] The upstream primer sequence was 5′-ATGATCCGAGATCCAATG-3′;

[0044] The downstream primer sequence was 5′-GTTACCCAAATGTGATGC-3′.

[0045] During primer design, to enhance the applicability and sensitivity of the primers, the designed primers are between 18 and 25 bp in length, and the primers do not interfere with each other. The above two primers can be obtained by artificial synthesis.

[0046] This embodiment also provides a kit for identifying whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low. The kit includes the above primer set.

[0047] The kit also includes common reagents for PCR amplification, such as 2× Rapid Taq Master Mix and ddH2O.

[0048] The primer set designed by the present invention has strong specificity and can accurately amplify the sequence carrying the SV molecular marker of the present invention.

[0049] By using the primer set or the kit of the present invention, the high or low content of quercetin 7-O-β-D-glucuronic acid in cassava can be accurately and efficiently predicted during the cassava seedling stage.

[0050] Example 3 Method for Identifying High or Low Content of Quercetin 7-O-β-D-Glucuronic Acid in Cassava

[0051] This example provides a method for identifying whether the content of cassava quercetin 7-O-β-D-glucuronic acid is high or low, and the specific method comprises the following steps:

[0052] S1. Extract genomic DNA from the cassava to be tested (cassava seedling stage is sufficient) and use it as a PCR template;

[0053] S2. Using the primer set in Example 2 and cassava genomic DNA as a template, PCR amplification of the SV molecular marker was performed;

[0054] The PCR amplification reaction system consisted of 12.5 μL of 2× Rapid Taq Master Mix, 1 μL of a 10 μM upstream primer, 1 μL of a 10 μM downstream primer, 1 μL of cassava genomic DNA, and 9.5 μL of ddH₂O. The amplification buffer (mix) was purchased from Novozymes Biotech Co., Ltd., and the primer set was synthesized by Beijing Aoke Dingsheng Biotechnology Co., Ltd.

[0055] The reaction program of PCR amplification was as follows: 95°C for 5 min; 95°C for 30 s, 60°C for 30 s, and 72°C for 30 s, 35 cycles; and 72°C for 5 min.

[0056] S3. Sequence the obtained PCR amplification product to determine whether it carries the SV molecular marker fragment.

[0057] When the nucleotide sequence of the PCR amplification product is as shown in SEQ ID NO: 1, and is 1565 bp in length, and the sequence is a DNA fragment represented by the SV molecular marker, that is, a sequence within the coordinate range of 1 to 1565 from the 5' end of the SV molecular marker, then the cassava to be tested carries the SV molecular marker, and the cassava to be tested has a high quercetin 7-O-β-D-glucuronic acid content, and is a cassava with a high quercetin 7-O-β-D-glucuronic acid content.

[0058] Otherwise, the cassava to be tested lacks the SV molecular marker, that is, a large fragment deletion occurs at bases 2657774 to 2659338 of chromosome 14 in the sample genome, and no amplification product is obtained by PCR amplification. In this case, the quercetin 7-O-β-D-glucuronic acid content of the cassava to be tested is low, and the cassava is low in quercetin 7-O-β-D-glucuronic acid content.

[0059] Alternatively, the PCR amplification product is subjected to electrophoresis detection, and the electrophoresis detection results are interpreted;

[0060] When only a 1565 bp band appears in the electrophoresis detection result, the cassava to be tested carries the SV molecular marker, and the quercetin 7-O-β-D-glucuronic acid content of the cassava to be tested is high, and the cassava is high in quercetin 7-O-β-D-glucuronic acid content;

[0061] Otherwise, when the electrophoresis detection result shows no band, the cassava to be tested lacks the SV molecular marker, and the quercetin 7-O-β-D-glucuronic acid content of the cassava to be tested is low, namely, the cassava with low quercetin 7-O-β-D-glucuronic acid content.

[0062] Among them, the relative content of quercetin 7-O-β-D-glucuronic acid in cassava with high quercetin 7-O-β-D-glucuronic acid content is generally ≥10416;

[0063] Low quercetin 7-O-β-D-glucuronic acid content The relative content of quercetin 7-O-β-D-glucuronic acid in cassava is generally <10416.

[0064] Example 4 Detection and Verification of SV Molecular Markers in Cassava

[0065] To verify the practicality of this SV molecular marker, several cassava plants (excluding the 337 cassava samples used for SV molecular marker development) were randomly selected in the cassava-growing area of Danzhou City, Hainan Province, China (109.5° east longitude, 19.5° north latitude). PCR amplification was performed using the method described in Example 3 to determine whether these plants carried the SV molecular marker. The relative content of quercetin 7-O-β-D-glucuronic acid in these cassava samples was also measured. The sequencing results are shown in Tables 1 and 2 below.

[0066] Table 1 Whether 20 cassava plants carry SV molecular markers and the relative content of quercetin 7-O-β-D-glucuronic acid

[0067]

[0068] Table 2 Statistics of whether 20 cassava plants carry SV molecular markers and relative contents of quercetin 7-O-β-D-glucuronic acid

[0069]

[0070] As can be seen from Tables 1 and 2, the method of the present invention can identify that the cassava carrying the SV molecular marker has a relatively high quercetin 7-O-β-D-glucuronic acid content, which is a cassava with a high quercetin 7-O-β-D-glucuronic acid content; the cassava lacking the SV molecular marker has a relatively low quercetin 7-O-β-D-glucuronic acid content, which is a cassava with a low quercetin 7-O-β-D-glucuronic acid content, indicating that the method of the present invention can be used to identify whether the cassava has a high or low quercetin 7-O-β-D-glucuronic acid content.

[0071] Example 5 Detection of SV Molecular Markers in Cassava

[0072] Furthermore, 207 cassava samples were typed using the method in Example 3. The results are as follows: Figure 2 As shown in the figure, 113 samples carried the SV molecular marker, while 94 samples lacked the SV molecular marker. The relative content levels of quercetin 7-O-β-D-glucuronic acid in these samples were tested and a T-test was performed. The results showed that the P value was 0.000001, indicating that the relative content of quercetin 7-O-β-D-glucuronic acid in cassava carrying the SV molecular marker was higher than that in cassava lacking the SV molecular marker, and the difference was significant.

[0073] All other parts not described in detail are prior art. Although the above embodiment provides a detailed description of the present invention, it is only a part of the embodiments of the present invention, not all of the embodiments. Those skilled in the art can also derive other embodiments based on this embodiment without inventiveness, and these embodiments all fall within the scope of protection of the present invention.

Claims

1. A molecular marker for high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease, characterized in that: The molecular marker is an SV molecular marker, and the sequence of the SV molecular marker is shown in SEQ ID NO: 1; The quercetin 7-O-β-D-glucuronic acid content of the cassava carrying the SV molecular marker is greater than the quercetin 7-O-β-D-glucuronic acid content of the cassava lacking the SV molecular marker.

2. A use of a reagent for detecting the molecular marker of high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease according to claim 1, characterized in that: The application is to detect whether cassava carries SV molecular markers by using the reagent to predict whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low; The reagent is used to detect whether cassava carries SV molecular markers; The quercetin 7-O-β-D-glucuronic acid content of the cassava carrying the SV molecular marker is greater than the quercetin 7-O-β-D-glucuronic acid content of the cassava lacking the SV molecular marker.

3. A primer set for detecting the molecular marker of high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root-knot nematode disease according to claim 1, characterized in that: The primer set includes: The upstream primer sequence is shown in SEQ ID NO: 2, specifically 5′-ATGATCCGAGATCCAATG-3′; The downstream primer sequence is shown in SEQ ID NO: 3, specifically 5′-GTTACCCAAATGTGATGC-3′; The primer set is used to detect whether cassava carries SV molecular markers; The quercetin 7-O-β-D-glucuronic acid content of the cassava carrying the SV molecular marker is greater than the quercetin 7-O-β-D-glucuronic acid content of the cassava lacking the SV molecular marker.

4. Use of a kit comprising the primer set according to claim 3, characterized in that: The kit is used to detect high quercetin 7-O-β-D-glucuronic acid content associated with resistance to root knot nematode disease; The quercetin 7-O-β-D-glucuronic acid content of the cassava carrying the SV molecular marker is greater than the quercetin 7-O-β-D-glucuronic acid content of the cassava lacking the SV molecular marker.

5. Use of the kit according to claim 4, characterized in that, The kit includes reagents for PCR amplification.

6. Use of the kit according to claim 5, characterized in that, The kit includes: 2× Rapid Taq Master Mix and ddH2O.

7. A method for identifying whether the content of cassava quercetin 7-O-β-D-glucuronic acid is high or low, characterized in that: The method comprises using the primer set of claim 3 and cassava genomic DNA as a template to perform PCR amplification on the SV molecular marker, sequencing the PCR amplification product, and determining whether the SV molecular marker is carried to predict whether the cassava has a high or low quercetin 7-O-β-D-glucuronic acid content; Alternatively, the kit according to any one of claims 4 to 6 is used to use cassava genomic DNA as a template to perform PCR amplification on the SV molecular marker, and the PCR amplification product is sequenced to determine whether it carries the SV molecular marker, so as to predict whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low; When the nucleotide sequence of the PCR amplification product is as shown in SEQ ID NO: 1, and the length is 1565 bp, the sequence is a DNA fragment represented by the SV molecular marker, and the cassava to be tested is cassava with a high quercetin 7-O-β-D-glucuronic acid content; Otherwise, the cassava to be tested lacks the SV molecular marker and no amplification product is obtained by PCR amplification, and the cassava to be tested is a cassava with low quercetin 7-O-β-D-glucuronic acid content.

8. The method for identifying whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low according to claim 7, characterized in that: The PCR amplification system includes: 2× Rapid Taq Master Mix, the upstream primer, the downstream primer, cassava genomic DNA and ddH2O; The reaction program of PCR amplification was as follows: 95°C for 5 min; 95°C for 30 s, 60°C for 30 s, and 72°C for 30 s, 35 cycles; and 72°C for 5 min.

9. A method for identifying whether the content of cassava quercetin 7-O-β-D-glucuronic acid is high or low, characterized in that: The method comprises using the primer set of claim 3 and cassava genomic DNA as a template to perform PCR amplification on the SV molecular marker, performing electrophoresis detection on the PCR amplification product, and interpreting the electrophoresis detection result to determine whether the SV molecular marker is carried, so as to predict whether the cassava has a high or low quercetin 7-O-β-D-glucuronic acid content; Alternatively, the kit according to any one of claims 4 to 6 is used to perform PCR amplification on the SV molecular marker using cassava genomic DNA as a template, the PCR amplification product is subjected to electrophoresis detection, and the electrophoresis detection result is interpreted to determine whether the SV molecular marker is carried, so as to predict whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low; When only a 1565 bp band appears in the electrophoresis detection result, the cassava carries the SV molecular marker and the cassava has a high quercetin 7-O-β-D-glucuronic acid content; When there is no band in the electrophoresis detection result, the cassava lacks the SV molecular marker and the cassava is low in quercetin 7-O-β-D-glucuronic acid content.

10. The method for identifying whether the cassava quercetin 7-O-β-D-glucuronic acid content is high or low according to claim 9, characterized in that: The PCR amplification system includes: 2× Rapid Taq Master Mix, the upstream primer, the downstream primer, cassava genomic DNA and ddH2O; The reaction program of PCR amplification was as follows: 95°C for 5 min; 95°C for 30 s, 60°C for 30 s, and 72°C for 30 s, 35 cycles; and 72°C for 5 min.

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