Application of genes, proteins and molecular markers related to edible rate of pumpkin fruit

Abstract

The present invention discloses the application of genes, proteins and molecular markers related to the edible rate of pumpkin fruits. The gene is a pumpkin CmoSYP71 gene, whose nucleotide sequence is shown in SEQ ID NO:1, and the amino acid sequence of the protein encoded by the gene is shown in SEQ ID NO:2, which can be used to detect the edible rate trait of pumpkin fruits or pumpkin auxiliary breeding. The present invention determines that the pumpkin CmoSYP71 gene is closely related to the edible rate trait of the fruit, and develops a molecular marker based on a G / C base mutation in the promoter region of the gene, which can be used to identify or screen the edible rate of pumpkin fruits, providing an important molecular basis for the breeding of new pumpkin varieties.

Description

Technical Field

[0001] The invention belongs to the technical field of crop genetic breeding, and in particular to an application of a CmoSYP71 gene, a protein and a molecular marker related to the edible rate of pumpkin fruit. Background Art

[0002] Pumpkin (Cucurbit moschata) is one of the most important melon crops in the world. Its fruit is rich in carotenoids, proteins, unsaturated fatty acids and amino acids, and is deeply loved by consumers. Pumpkin fruit has a variety of traits, and the edible rate of the fruit (the ratio of the flesh area of ​​the longitudinal section of the fruit to the area of ​​the whole fruit) is an important quality trait of pumpkin that is not visible inside. The edible rate of pumpkin fruit is calculated based on the two-dimensional area ratio, which can reflect the overall quality of the fruit. Increasing the edible rate can effectively increase the yield and commerciality, and reduce the transportation and storage costs.

[0003] Previous studies have shown that the edible rate of pumpkin fruit is basically determined during the young fruit stage, and cell division is an important factor affecting the edible rate of pumpkin fruit. Further exploration and utilization of genes, proteins and molecular markers related to the edible rate of pumpkin fruit will be of great significance to promoting the creation of new materials related to the edible rate of pumpkin fruit and the breeding of new varieties, and accelerating the molecular breeding process of pumpkin. Summary of the invention

[0004] Based on this, the purpose of the present invention is to provide an application of CmoSYP71 gene, protein, and molecular markers related to the edible rate of pumpkin fruit. The pumpkin CmoSYP71 gene is involved in cell division and is a regulatory gene for the edible rate trait of pumpkin fruit.

[0005] The specific technical solutions for achieving the above-mentioned invention objectives include the following.

[0006] In a first aspect of the present invention, a use of the pumpkin CmoSYP71 gene in detecting pumpkin fruit edible rate traits or pumpkin assisted breeding is provided, and the nucleotide sequence of the pumpkin CmoSYP71 gene is shown in SEQ ID NO:1.

[0007] The second aspect of the present invention provides the use of the protein encoded by the pumpkin CmoSYP71 gene in detecting the edible rate trait of pumpkin fruit or pumpkin assisted breeding, and the amino acid sequence of the encoded protein is shown in SEQ ID NO:2.

[0008] The third aspect of the present invention provides an application of a molecular marker in detecting the edible rate trait of pumpkin fruit or pumpkin assisted breeding, wherein the molecular marker is located at position 865631 of chromosome 14 of the pumpkin genome, where the base is G or C.

[0009] In the fourth aspect of the present invention, a molecular marker amplification primer pair is provided, comprising an upstream primer whose sequence is shown in SEQ ID NO:3, and downstream primers whose sequences are shown in SEQ ID NO:4 and SEQ ID NO:5; the molecular marker is located at position 865631 of chromosome 14 of the pumpkin genome, where the base is G or C.

[0010] The fifth aspect of the present invention provides a kit for detecting the edible rate of pumpkin fruit or screening pumpkin varieties with high edible rate of fruit, wherein the kit comprises the above-mentioned amplification primers.

[0011] The sixth aspect of the present invention provides the use of the above-mentioned amplification primer pair or kit in detecting the edible rate trait of pumpkin fruit or pumpkin assisted breeding.

[0012] The seventh aspect of the present invention provides a method for detecting the edible rate trait of pumpkin fruit, comprising the following steps: using the pumpkin genome to be tested as a template, using the above-mentioned amplification primer pair to perform PCR amplification, and detecting whether the 865631 position of chromosome 14 of the pumpkin genome to be tested is G or C.

[0013] The eighth aspect of the present invention provides a method for identifying pumpkin varieties with high edible fruit rates and low edible fruit rates, comprising the following steps: using the pumpkin genome to be tested as a template, performing PCR amplification using the above-mentioned amplification primer pairs, and analyzing the genotype of the PCR amplification product.

[0014] The ninth aspect of the present invention provides a method for screening pumpkin varieties with high edible fruit rate, comprising the following steps: using the pumpkin genome to be tested as a template and using the above-mentioned amplification primer pair to perform PCR amplification.

[0015] In the present invention, based on the finding that "the edible rate of the fruits of the parental materials LA-3 and SK-2 has been determined at the young fruit stage, and the edible rate of the fruits of LA-3 is significantly higher than that of SK-2, and the difference in cell number is an important reason for the difference in the edible rate of the fruits of the two", the inventors used LA-3 and SK-2 to create an F2 segregation population, and through gene mapping, the main effect site of the fruit edible rate Cmofer14.1 was discovered on chromosome 14 of pumpkin. Only the CmoSYP71 gene in this site showed significant expression differences in the young fruits of the two parents, and CmoSYP71 was more highly expressed in the high fruit edible rate pumpkin LA-3. Gene annotation results show that this gene is involved in the positive regulation of cell division. Therefore, the CmoSYP71 gene is closely positively correlated with the edible rate trait of pumpkin fruit.

[0016] Furthermore, the inventors found that there was a C / G base variation in the promoter region of the CmoSYP71 gene (268 bp upstream of the start codon ATG, position 865631 of chromosome 14), which was C in the high fruit edible rate pumpkin LA-3 and G in the low fruit edible rate pumpkin SK-2. Based on the mutation site, a molecular marker was developed, and the F2 population genotype analysis showed that the mutation of this site to G significantly reduced the edible rate of pumpkin fruit, so that the molecular marker can be used to identify and / or screen the fruit edible rate trait of pumpkin, providing an important molecular basis for the breeding of new pumpkin varieties. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 It is a longitudinal section of the parent material and F1 generation fruit in Example 1 of the present invention.

[0018] Figure 2 This is the normal distribution PP diagram of the fruit edible rate of the F2 population in Example 1 of the present invention.

[0019] Figure 3 This is the result of the gene localization of the main QTL Cmofer14.1 of pumpkin fruit edible rate in Example 1 of the present invention. The horizontal axis is the position of the linkage group, the vertical axis represents the LOD value, the red dotted line represents the threshold of LOD (p value is less than 0.05), and the area above the dotted line represents a significant association.

[0020] Figure 4 This is the precise positioning of the pumpkin fruit edible rate gene in Example 1 of the present invention.

[0021] Figure 5 This is the result of gene expression analysis in the Cmofer14.1 localization interval in Example 1 of the present invention.

[0022] Figure 6 This is the genotyping result of the F2 genetic population using the primer pair of molecular markers in Example 2 of the present invention. DETAILED DESCRIPTION

[0023] In order to facilitate the understanding of the present invention, the present invention will be described more fully below. The present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the understanding of the disclosure of the present invention more thorough and comprehensive.

[0024] Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as those commonly understood by those skilled in the art to which the present invention belongs. The terms used in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The term "and / or" used in the present invention includes any and all combinations of one or more of the related listed items.

[0025] Unless otherwise specified, the examples are based on conventional experimental conditions, such as Sambrook et al. Molecular Cloning: a Laboratory Manual (Sambrook J & Russell DW, Molecular Cloning: a Laboratory Manual, 2013), or the conditions recommended by the manufacturer's instructions.

[0026] Based on the high-density genetic map and QTL positioning of pumpkin, the present invention excavated an important site Cmofer14.1 on chromosome 14 of the pumpkin genome that regulates the edible rate of pumpkin fruit, which can explain 22.5% of the phenotypic variation. The site contains 7 genes (Table 1). Expression analysis found that only CmoSYP71 in this interval was significantly differentially expressed in the young fruits of the two parental materials; the gene annotation results showed that CmoSYP71 was related to cell division; based on the results of parental resequencing, sequence variation analysis was performed, and it was found that there was a C / G base variation at the position 268bp upstream of the gene promoter. The genotype analysis of the F2 population showed that the mutation of this site to G significantly reduced the edible rate of pumpkin fruit. Based on the above analysis results, CmoSYP71 was determined to be a pumpkin fruit edible rate regulating gene, which can be used to breed new pumpkin varieties with high fruit edible rate.

[0027] In some embodiments of the present invention, the use of pumpkin CmoSYP71 gene in detecting pumpkin fruit edible rate trait or pumpkin assisted breeding is disclosed, and the nucleotide sequence of the pumpkin CmoSYP71 gene is shown in SEQ ID NO: 1. The CmoSYP71 gene is related to pumpkin fruit edible rate and can be used in molecular breeding related to pumpkin fruit edible rate.

[0028] In other embodiments of the present invention, disclosed is the use of the protein encoded by the pumpkin CmoSYP71 gene in detecting pumpkin fruit edible rate traits or pumpkin assisted breeding, and the amino acid sequence of the encoded protein is shown in SEQ ID NO:2.

[0029] In other embodiments of the present invention, a molecular marker is disclosed for use in detecting pumpkin fruit edible rate traits or pumpkin assisted breeding. The molecular marker is located at position 865631 of chromosome 14 of the pumpkin genome, where the base is G or C.

[0030] In other embodiments of the present invention, a molecular marker amplification primer pair is disclosed, comprising an upstream primer having a sequence as shown in SEQ ID NO:3, and downstream primers having sequences as shown in SEQ ID NO:4 and SEQ ID NO:5; the molecular marker is located at position 865631 of chromosome 14 of the pumpkin genome, where the base is G or C.

[0031] In some of the embodiments, the 5' end of the downstream primer whose sequence is shown in SEQ ID NO:4 is connected to a FAM group; the 5' end of the downstream primer whose sequence is shown in SEQ ID NO:5 is connected to a HEX group.

[0032] In other embodiments of the present invention, a kit for detecting the edible rate trait of pumpkin fruit or screening pumpkin fruit varieties with high edible rate is disclosed, and the kit comprises the above-mentioned amplification primer pair.

[0033] In other embodiments of the present invention, the use of the above-mentioned amplification primer pair or kit in detecting the edible rate trait of pumpkin fruit or pumpkin assisted breeding is disclosed.

[0034] In other embodiments of the present invention, a method for detecting the edible rate trait of pumpkin fruit is disclosed, comprising the following steps: using the pumpkin genome to be tested as a template, performing PCR amplification using the above-mentioned amplification primer pair, and detecting whether the 865631 position of chromosome 14 of the pumpkin genome to be tested is G or C.

[0035] In other embodiments of the present invention, a method for identifying pumpkin varieties with high edible fruit rates and low edible fruit rates is disclosed, comprising the following steps: using the pumpkin genome to be tested as a template, performing PCR amplification using the above-mentioned amplification primer pair, and analyzing the genotype of the PCR amplification product. Specifically, if the detected genotype is heterozygous G, it is a pumpkin variety with a medium to high edible fruit rate, and if it is homozygous G, the fruit edible rate is the lowest, and it is a pumpkin variety with a low edible fruit rate.

[0036] In other embodiments of the present invention, a method for screening pumpkin varieties with high edible fruit rate is disclosed, comprising the following steps: using the pumpkin genome to be tested as a template, and performing PCR amplification using the above-mentioned amplification primer pair.

[0037] The present invention is described in detail below with reference to the accompanying drawings and specific embodiments.

[0038] Example 1 Location and analysis of genes related to pumpkin fruit edible rate

[0039] 1. Experimental Materials

[0040] The high edible fruit rate pumpkin LA-3 and the low edible fruit rate pumpkin SK-2 are both core breeding materials of the Vegetable Research Institute of Guangdong Academy of Agricultural Sciences.

[0041] 2. Experimental steps

[0042] (1) The high edible fruit pumpkin LA-3 (fruit edible rate 73.5%) and the low edible fruit pumpkin SK-2 (fruit edible rate 49.9%) were used as parents, and the F1 (fruit edible rate 60.2%) was obtained by hybridization. The F2 segregating population was further created. The longitudinal section of the parent material and the F1 generation fruit is shown in Figure 1. Figure 1 shown.

[0043] (2) The edible rate of the fruit in the F2 population was investigated and SPSS software was used to test whether the trait conformed to the normal distribution. The results are as follows: Figure 2 The PP diagram (Probability-probability Plot) shows that the probability distribution of the variables is basically a straight line, and the edible rate of the fruit in the F2 population conforms to the normal distribution.

[0044] (3) A high-density genetic map of pumpkin was constructed using the F2 population. Combined with the phenotype, the main QTL locus Cmofer14.1 controlling the edible rate of the fruit (the ratio of the flesh area of ​​the longitudinal section of the fruit to the area of ​​the whole fruit) was found on chromosome 14 through gene mapping, which can explain 22.5% of the phenotypic variation ( Figure 3 ). Molecular markers (M8656-M8962) were designed inside and at both ends of the Cmofer14.1 positioning interval, and the fruit edible rate gene was precisely located to an interval of 30.6 kb. The results are as follows Figure 4 There are a total of 7 genes in the localization interval of this important site Cmofer14.1, and their annotation information is shown in Table 1.

[0045] Table 1 Gene annotation information within the QTLCmofer14.1 localization interval

[0046] GeneID NR annotation CmoCh14G001950 syntaxin-71-like [Cucurbita moschata] CmoCh14G001960 protein indeterminate-domain9-like [Cucurbita moschata] CmoCh14G001970 uncharacterized protein LOC111441966 [Cucurbita moschata] CmoCh14G001980 probable E3 ubiquitin-protein ligase LOG2 [Cucurbita moschata] CmoCh14G001990 V-type proton ATPase subunit G-like [Cucurbita moschata] CmoCh14G002000 unknown CmoCh14G002010 60S ribosomal protein L12-1-like [Cucurbita moschata]

[0047] (4) Analyze the expression of seven genes in the Cmofer14.1 localization interval in young fruits of parental materials, such as Figure 5 As shown, only CmoCh14G001950 was significantly differentially expressed between the parental materials, and was more highly expressed in pumpkins with high fruit edible rate. Gene annotation showed that CmoCh14G001950 (named CmoSYP71) was a Syntaxinfamilyprotein encoding gene, which may be involved in cell division, increase the number of cells, and thus improve the fruit edible rate.

[0048] (5) Based on the results of parental resequencing, further sequence variation analysis of the gene revealed a C / G base variation at 268 bp upstream of the CmoSYP71 gene promoter, which was C in the high edible fruit rate pumpkin LA-3 and G in the low edible fruit rate pumpkin SK-2.

[0049] Therefore, it was judged that the CmoSYP71 gene (nucleotide sequence as shown in SEQ ID NO: 1 (including 795 bp of the coding region and its upstream promoter sequence, i.e. 2000 bp upstream of the start codon ATG), and the encoded amino acid sequence as shown in SEQ ID NO: 2) was related to the edible rate of pumpkin fruit.

[0050] SEQ ID NO:1

[0051] atggtttggttgaacttataatatatatatatatatataattaaccaaataataaaattgtttaaaaa aataaaaaatttaaaataagtgtcaaaattttctcattgaatttcaattgcaaccgttctagcccaagactttagg cgtttcttcatgtcagaggtggaagaaacttttggcatccttagggataaaagaggggaatttggtcggtttattg tccatgcctctgaaagatatggtcttgctatatctcttttcttatggtcatcgttttcatctgcatcgtcgttgtc gtcgtcgtcgtcttcgggattttgtttgtccattcgagttttaaacctttgcaaaccgggaaccaacttcaccatt tgagggctcaaattttctgtcctaaattcaaaccctaaatccataaaccctttaacttcctccatctccaactccc ccaagcttttacaactcatagctttgttcaattttttcctcactgcacacaacaattcttaatcacatcaatataa catgaatatacgttcctctccacattttattaattgggtttcaaagtttatgtaggatactaataataattcaact cacttgtaaattcaggatcagtgagactttcttccattgcctcttcctttccatcttgtttatatttgtctccatc gttgctatcatcttcctgttcaaaacaaacatcaaaatcactttttatattatatcaaagcacgagagatcgaacg ttcaaaactgtcgtcgttcttactcgttatcatatcctatcgagcacagggagaagggttacctgggagacgaggg aggtgtcgtcgaggaagatgcggtcggaggaaagggaagaagggaagtttgagaaaggaaaagttgggtgaagatg ggaaggggaaggatgtggagaggaaaaatgatttggtggtgccagaggcgatccatggctgaaacaagacataga gtgtttgtgttggtgatagccatggaagagcaaagagtggaagggagacaaagattgggatataaaagaagccatg gaagtgaacacagagttgtggtggaaatgaaagaaggaatgagttctcaaccccaatcaaaaattaaaggaat attggaggatgtggccataagtgaatatcaatccatatcctttctcttttttctttttctttttccataaccaaatt actcttttacccccatagtttacgtataaagccggtttttttaataaaaaattattctctatcagtactcaacttga attgatttgattaagtctgtgatagtaatgactcgatattacaaaatctttaaaatttttattttatacgttacta cgtatgaatttttaaaatttctttaaaaatcatatttgttagggaaaatatttaattattcaatttttaattaggt tacaatcaacaaatgggcatttggtctagtggtatgattctcgcttagggtgcgagaggtcccgagttcaattctc ggaatgccccaaatcaatcaaccttttctttccattttgggccttcatggtaacttccggcctgaaaaatgaaatg ggccactttgctgtgggcttcacaattttgggccgcccctctcgacctgaaattagaggataaaagggaattgtta tttgtttttatatgaaattttagatttattgtttaatcgggtgttaattaatggggtagtttttcataCcaaattc cacgattttgttgatttttaaaattagacatataaatggaagcttaaaatttgtgagctttttttgtttgtttgtt ttgtgacaaaatagtcgtaatttatcataaatttaaagcttttaaactgaaagtttgcctcggtgagtagaacaat cagcggcggtactggcactggcgcacaacttttgtagatattcggaaccgctttcaccgttgtagctcaccggaaa ccctatcgaagcacggaggcgaagaatcgaagATGAGCGTGATCGACCTTTTGACCAGAGTAGACGCGATCTGCCAGAAATACGACAAATATGATGTAGAGAAGCAGAGAGATCTCAATGTCTCTGGTGACGATGCTTTCGCTCGACTCTATGCCACCGTCGAAGCCGACATTGAAGCCGCTCTCCAGAAAGCGGAGGATGCTTCCAGAGAGAAGAATAGGGCATCGGTTG TGGCGTTGAATGCGGAGATTCGTCATACCAAGGCTCGGTTACTAGAGGAGGTCCCCAAGTTGCAGAGATTAGCTGTAAAGAGGGTTAAAGGACTATCAACTGAAGATCTTACCACTCGAAATGATTTGGTGCTTGCATTGCCGGATAGGATTCAAGCTATACCAGATGGAACTGCCTCTTCCTCAAAGAAGAATGGGGGT TGGACCTCCTCAACTTCACGTGCTGAAATCAAATTTGACTCGGATGGGCGATTTGATAATGAGTACTTCCAACACACTGAGGAGTCGAGTCAATTCAGACAAGAGTATGAAATGCGGAAAATGAAGCAGGATCAAGGATTAGACATGATATCAGAGGGGGTTGGATACTCTGAAGAACATGGCGCATGATATGAATGAGG AAATAGACAGGCAAGTTTCCTTTGATGGACGAGATTGACACTAAGGTGGACAAGGCTGCATCTGACCTTAAGAACACTAACGTTAGATTAAGGGACACAGTTAACCAGCTAAGGTCCAGCAGAAATTTCTGCATCGATATCATTTTGTTGTGTATAATCTTGGGTATTGCTGCCTATCTATACAATGTGTTGAAGAAGTGA

[0052] SEQ ID NO:2

[0053] MSVIDLLTRVDAICQKYDKYDVEKQRDLNVSGDDAFARLYATVEADIEAALQKAEDASREKNRASVVALNAEIRHTKARLLEEVPKLQRLAVKRVKGLSTEDLTTRNDLVLALPDRIQAIPDGTASSSKKNG GWTSTSRAEIKFDSDGRFDNEYFQHTEESSQFRQEYEMRKMKQDQGLDMISEGLDTLKNMAHDMNEEIDRQVPLMDEIDTKVDKAASDLKNTNVRLRDTVNQLRSSRNFCIDIILLCIILGIAAYLYNVLKK

[0054] Example 2 Molecular markers associated with pumpkin fruit edible rate and their verification

[0055] The following steps are involved:

[0056] 1. According to the SNP site in Example 1 (i.e., the C / G base variation at 268 bp upstream of the CmoSYP71 gene promoter), the amplification primers of the molecular marker were designed using primerpremier 5.0 software, including an upstream primer F, a downstream primer Rc and a downstream primer Rg, the 5' end of the downstream primer was connected with a fluorescent label sequence, wherein the 5' end of the downstream primer Rc was connected with a fluorescent label sequence 5'-GAAGGTGACCAAGTTCATGCT-3' with a FAM group, and the 5' end of the downstream primer Rg was connected with a fluorescent label sequence 5'-GAAGGTCGGAGTCAACGGATT-3' with a HEX group; the bold bases C (SEQ ID NO: 4) and G (SEQ ID NO: 5) at the 3' end were typing sites.

[0057] Upstream primer F (SEQ ID NO: 3):

[0058] CACTTTGCTGTGGGCTTCAC

[0059] Downstream primer Rc (SEQ ID NO: 4):

[0060] GAAGGTGACCAAGTTCATGCT CAACAAAATCGTGGAATTTGC

[0061] Downstream primer Rg (SEQ ID NO: 5):

[0062] GAAGGTCGGAGTCAACGGATT CAACAAAATCGTGGAATTTGG

[0063] 2. Take the young leaves of the F2 population sample, obtain the pumpkin genomic DNA to be tested by CTAB extraction method, and use the above primer set for PCR amplification. The PCR reaction system is 10μL, which contains 2×PARMS mastermix 5μL, upstream primer 0.15μL, downstream primer Rc 0.15μL, downstream primer Rg 0.15μL, DNA (10-100ng) 1μL, ddH2O 3.55μL. The PCR reaction program is: 94℃, 15min; 94℃, 20s, 65℃-57℃, 1min, 10 cycles; 94℃, 20s, 57℃, 1min, 30 cycles.

[0064] 3. After the PCR reaction is completed, use an ELISA reader with 3 channels to read the fluorescence signal. The fluorescence signal is combined with the label information to obtain the genotyping diagram, and the genotype results are output according to the different colors. The samples aggregated on the X-axis and with blue fluorescence signals are genotype FAM, with a base of GG; the samples aggregated on the Y-axis and with green fluorescence signals are genotype HEX, with a base of CC; the samples aggregated in the middle and with red fluorescence signals are heterozygous genotypes, with a base of GC. The test results are shown in Figure 6 .

[0065] 58 materials with obvious phenotypes were selected from the F2 population (including 2 materials with high fruit edible rate, 35 materials with medium and high fruit edible rate, and 17 materials with low fruit edible rate). Combined with genotype analysis, it was found that the mutation from C to G at this site significantly reduced the edible rate of pumpkin fruit (the genotypes of the 2 materials with high fruit edible rate were CC, the average fruit edible rate was 71.7%, and the coincidence rate was 100%; the genotypes of the 35 materials with medium and high fruit edible rate were GC, the average fruit edible rate was 63.2%, and the coincidence rate was 100%; the genotypes of the 14 materials with low fruit edible rate were GG, the average fruit edible rate was 53.21%, and the coincidence rate was 82.3%; the overall average coincidence rate of the 58 materials was 94%). Therefore, the variation of this site is closely related to the edible rate of fruit. According to the molecular markers of this variation site, it can be used to detect the edible rate trait of pumpkin fruit.

[0066] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0067] The above-mentioned embodiments only express several implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. An application of a molecular marker in detecting the edible rate trait of pumpkin fruit, wherein the molecular marker is located at the 1733 bp position of the nucleotide sequence shown in SEQ ID NO: 1, and the base at this position is C or G.

2. A pair of primers for amplification of molecular markers, characterized in that: It comprises an upstream primer with a sequence as shown in SEQ ID NO:3, and downstream primers with sequences as shown in SEQ ID NO:4 and SEQ ID NO:5; the molecular marker is located at the 1733 bp position of the nucleotide sequence as shown in SEQ ID NO:1, and the base at this position is G or C.

3. The amplification primer pair according to claim 2, characterized in that: The 5' end of the downstream primer whose sequence is shown in SEQ ID NO:4 is connected to a FAM group; the 5' end of the downstream primer whose sequence is shown in SEQ ID NO:5 is connected to a HEX group.

4. A kit for detecting the edible rate of pumpkin fruit or screening pumpkin varieties with high edible rate of fruit, characterized in that: The kit comprises the amplification primer pair according to claim 2 or 3.

5. Use of the amplification primer pair according to claim 2 or 3, or the kit according to claim 4, in detecting the edible rate trait of pumpkin fruit.

6. A method for detecting the edible rate of pumpkin fruit, characterized in that: The following steps are involved: The pumpkin genome to be tested is used as a template, and PCR amplification is performed using the amplification primer pair described in claim 2 or 3 to detect whether the 1733bp position of the pumpkin CmoSYP71 gene to be tested is G or C. The nucleotide sequence of the CmoSYP71 gene is shown in SEQ ID NO:

1.

7. A method for identifying pumpkin varieties with high edible fruit rates and pumpkin varieties with low edible fruit rates, characterized in that: The following steps are involved: The pumpkin genome to be tested is used as a template, PCR amplification is performed using the amplification primer pair described in claim 2 or 3, and the genotype of the PCR amplification product is analyzed.

8. A method for screening pumpkin varieties with high edible fruit rate, characterized in that: The following steps are involved: The pumpkin genome to be tested is used as a template and PCR amplification is performed using the amplification primer pair described in claim 2 or 3.

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