A method for constructing SSR marker primers and fingerprints of ulmus aurantium germplasm resources

Abstract

The present invention belongs to the technical field of molecular biology, and is specifically a method for constructing an SSR marker primer and fingerprint map of eriophora pubescens germplasm resources, wherein the SSR marker primer group comprises a primer group having sequences such as SEQ ID NO.1-2, SEQ ID NO.3-4, SEQ ID NO.5-6, SEQ ID NO.7-8, SEQ ID NO.9-10, SEQ ID NO.11-12, SEQ ID NO.13-14, SEQ ID NO.15-16, SEQ ID NO.17-18, and SEQ ID NO.19-20. The present invention uses SSR primers to reveal the genetic differences and kinship of different eriophora pubescens germplasms, and provides a reference for the identification, development, protection and utilization of eriophora pubescens germplasm resources.

Description

Technical Field

[0001] The invention relates to the technical field of molecular biology, and in particular to a method for constructing an SSR marker primer and a fingerprint map of an ulmus auricula germplasm resource. Background Art

[0002] As a kind of precious wild medicinal and edible dual-purpose fungi unique to Northeast my country, Gloeostereumincarnatum S.Ito et Imai has a fruiting body rich in multivitamins, trace elements and amino acids necessary for the human body, rich in nutrition and has multiple pharmacological activities. Due to the lack of unified management and effective identification methods, there is confusion in the strain management and identification of Gloeostereumincarnatum, and the phenomenon of the same species with different names and the same name with different species is very common, which has a serious impact on the sustainable development of the industry. At present, there are not many cultivars used in production, and the existing cultivars are mostly directly applied to production after being separated from wild Gloeostereumincarnatum tissues. The strain degradation phenomenon is serious, and the output and quality are very unstable. It is impossible to determine which characters can be stably inherited, and there is no standard for distinguishing the character differences between varieties, which is not conducive to the development of germplasm resources of Gloeostereumincarnatum and the research on new variety breeding. Therefore, it is urgent to develop a method with strong operability and accurate identification of Gloeostereumincarnatum germplasm (variety) strains.

[0003] Germplasm resource evaluation is the basis for the utilization of edible fungi germplasm resources, and is mainly carried out from four aspects: morphology, cytology, biochemistry, and molecule. At present, the evaluation of elm ear germplasm resources is mainly based on agronomic traits (morphology), but germplasms with similar phenotypes are difficult to distinguish, and the phenotypic traits are greatly affected by the cultivation environment, making it difficult to accurately reflect the true genetic relationship. The SSR molecular markers developed based on the elm ear genome are abundant in number, evenly distributed, highly polymorphic, and have good repeatability. They are an ideal molecular marker technology and have been widely used in variety identification of field crops, horticultural crops, and database establishment. They have also been selected as the preferred marker for constructing a DNA fingerprint database. DNA fingerprinting is a powerful tool for identifying varieties, with the advantages of being fast and accurate, and has also been widely used in the diversity and purity identification of variety resources of many crops. The construction of the SSR molecular marker fingerprint map of elm ear germplasm resources has not been reported, which has seriously hindered the development of elm ear germplasm resources and the research on new variety breeding. Summary of the invention

[0004] In order to solve the above technical problems, the present invention provides a method for constructing SSR marker primers and fingerprints of ulmus auricula germplasm resources.

[0005] A combination of SSR marker primers for ulmus fusca germplasm resources, the SSR marker primer group comprising primer group one whose sequences are respectively as described in SEQ ID NO.1-2, primer group two as described in SEQ ID NO.3-4, primer group three as described in SEQ ID NO.5-6, primer group four as described in SEQ ID NO.7-8, primer group five as described in SEQ ID NO.9-10, primer group six as described in SEQ ID NO.11-12, primer group seven as described in SEQ ID NO.13-14, primer group eight as described in SEQ ID NO.15-16, primer group nine as described in SEQ ID NO.17-18, and primer group ten as described in SEQ ID NO.19-20.

[0006] A kit comprises the above-mentioned Auricularia auricularia germplasm resource SSR marker primer combination.

[0007] The application of the described ulmus fusilli germplasm resource SSR marker primer combination or the described kit in constructing a fingerprint map of ulmus fusilli germplasm resources.

[0008] The application of the described ulmus fusilli germplasm resource SSR marker primer combination or the described kit in identifying ulmus fusilli germplasm resources.

[0009] A method for constructing a fingerprint map of elm ear germplasm resources, comprising the following steps: using the elm ear germplasm resource SSR marker primer combination described in claim 1 to perform PCR amplification on the DNA of the elm ear germplasm resources, screening core primers, reading band size data, screening to obtain polymorphic fragments, converting the polymorphic fragments into 0, 1 matrix data in a binary manner, and obtaining a molecular ID card of the elm ear germplasm resources.

[0010] Preferably, the reaction system for PCR amplification is 5 μL of dNTP Mixture, 0.8±0.1 μL of forward primer, 0.8±0.1 μL of reverse primer, 1±0.1 μL of DNA, and ultrapure water added to 11 μL.

[0011] Preferably, the reaction conditions for the PCR amplification are pre-denaturation at 94°C for 5 min; denaturation at 94°C for 45 s, annealing at 60°C for 45 s, extension at 72°C for 1 min, for a total of 35 cycles; and extension at 72°C for 10 min.

[0012] The application of the construction method in identifying Auricularia auricularia germplasm resources.

[0013] A method for identifying eriophora germplasm resources, comprising the following steps: using the eriophora germplasm resource SSR marker primer combination described in claim 1 to perform PCR amplification on the eriophora germplasm resource DNA to be tested, comparing the obtained PCR amplification band with the eriophora germplasm resource fingerprint map constructed by the construction method, and identifying the type of the eriophora germplasm resource.

[0014] Compared with the prior art, the present invention has the following beneficial effects:

[0015] The 10 pairs of core SSR primers of the present invention can be used to compare different elm ear germplasm resources, and the establishment of SSR molecular fingerprints can accurately detect target genes. The present invention uses SSR markers to study the genetic diversity and draw fingerprints of elm ear germplasm resources, revealing the genetic differences and kinship of different elm ear germplasms, and providing a reference for the identification, development, protection and utilization of elm ear variety resources. The operation is simple, sensitive and timely, and the constructed resource fingerprint is of great significance for the identification of rare edible elm ear varieties in my country, the breeding of new varieties, and the efficient utilization and protection of resources. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 is the allele distribution map of 29 primer pairs;

[0017] Figure 2 This is a cluster analysis diagram of Ulmus auricularia strains based on SSR amplification. Note: The two branch colors are the results of population structure division, pink: A, blue: B; the four font colors are the results of cluster division based on agronomic traits, group I: red, group II: yellow, group III: blue, group IV: green;

[0018] Figure 3 The QR code fingerprint pattern and scanning results of the GH93 strain;

[0019] Figure 448, GH49, GH50, GH51, GH52, GH53, GH56, GH59, GH61, GH62, GH63, GH64, GH66, GH67, GH68, GH70, GH71, GH72, GH73, GH74, GH76, GH78, GH80, GH83, GH85, GH87, GH89, GH90, GH91, GH92, GH93, GH94, GH96, GH97, GH98, GH99, GH100, GH101, GH102, GH103, GH104, GH105, GH106, GH107, GH108, GH109, ​​GH110 0, GH91, GH92, GH93, GH94, GH95, GH96, GH100, GH104, GH105, GH106; Figure B shows a total of 41 columns of results, the first column is the result of Marker; the test strain numbers in columns 2-41 are GH110, GH112, GH113, GZ3, GZ12, GZ13, GZ26, GZ31, GZ32, GZ33, GZ Results for 46, GZ47, GZ48, GZ58, GZ64, GZ68, GZ69, GZ70, GZ71, GZ72, GZ77, GZ78, GZ80, GZ85, GZ86, GZ87, GH89, GZ90, GZ94, GZ95, GZ99, GZ102, GZ104, GZ106, GZ109, GZ111, GZ113, C, D, 126-15;

[0020] Figure 5It is the electrophoresis diagram of SSR500 primer amplification. In Figure A, there are 62 columns of results. The 1st and 62nd columns are the results of Marker; the test strains in columns 2-61 are numbered GH1, GH2, GH3, GH4, GH10, GH14, GH16, GH17, GH20, GH21, GH23, GH25, GH27, GH31, GH32, GH33, GH34, GH37, GH38, GH41, GH43, GH45, GH46, GH47, GH48, GH49, GH50, GH51, GH52, GH53, GH54, GH55, GH56, GH57, GH58, GH59, GH60, GH61, GH62, GH63, GH64, GH65, GH66, GH67, GH68, GH69, GH70, GH71, GH72, GH73, GH74, GH75 H48, GH49, GH50, GH51, GH52, GH53, GH56, GH59, GH61, GH62, GH63, GH64, GH66, GH67, GH68, GH70, GH71, GH7 2. GH73, GH74, GH76, GH78, GH80, GH83, GH85, GH87, GH90, GH91, GH92, GH93, GH94, GH95, GH96, GH100, GH10 4. Results of GH105 and GH106; Figure B shows a total of 41 columns of results, the first column is the result of Marker; the test strain numbers in columns 2-41 are GH110, GH112, GH113, GZ3, GZ12, GZ13, GZ26, GZ31, GZ32, GZ33, GZ46, GZ47, GZ48, GZ58, GZ64, GZ68, GZ69, GZ70, GZ71, GZ72, GZ77, GZ78, GZ80 , GZ85, GZ86, GZ87, GH89, GZ90, GZ94, GZ95, GZ99, GZ102, GZ104, GZ106, GZ109, GZ111, GZ113, C, D, and 126-15; Figure C shows a total of 10 columns of results, the 1st and 10th columns are the results of Marker; the 2nd to 9th columns are the results of the test strains numbered GH1, GH3, GH10, GH51, GH59, GZ69, GZ72, and GZ78. DETAILED DESCRIPTION

[0021] The specific embodiments of the present invention are described in detail below, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the protection scope of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified.

[0022] Example 1

[0023] Analysis on SSR Genetic Diversity and Construction of Molecular Fingerprint of Auricularia auricularia A ...

[0024] 1. Test strains

[0025] In this experiment, 100 strains of elm ear were used as test materials, including 2 parent strains, 1 hybrid strain, 33 self-pollinating strains and 64 backcross strains. The 2 parent strains were parent C and parent D, parent C was an early-maturing medium-yield strain, parent D was a late-maturing high-yield strain, and 1 hybrid strain was 126-15, 126-15 was a medium-maturing medium-yield strain, and the hybrid strain 126-15 was obtained by hybridizing parent C and parent D; the monokaryotic strain of 126-15 was self-pollinated to obtain the self-pollinating strain; the monokaryotic strain of 126-15 was backcrossed with C and D to obtain the backcross strain. All the test strains have completed phenotypic diversity analysis and are stored in the College of Horticulture of Jilin Agricultural University. The strain information is shown in Table 1.

[0026] Table 1 Test strain numbers

[0027] Accession number Strain number Strain type Strain source C GY027 wild strain Jilin Province Baishan City Sandaogou State Forest Farm D yrzp-1 Cultivated strains Yehe Town, Siping City, Jilin Province 126-15 Hybrid strains Hybrid strain of GY027 and yrzp-1 GZ3~GZ113 selfing strain Self-fertilized strain 126-15 GH1~GH61 backcross strain 126-15 is a result of backcrossing with parent D GH62~GH113 backcross strains 126-15 is a result of backcrossing with parent C

[0028] 2. Test methods

[0029] (1) Extraction of Ulmus auricula DNA

[0030] The test strain was inoculated on a PDA medium covered with cellophane and cultured at 25°C in the dark for 8-11 days. The mycelium was scraped off with a small spoon and collected in a mortar. Liquid nitrogen was quickly poured into the mycelium to grind it into powder. About 0.2 g was weighed and placed in a sterilized 2 ml centrifuge tube and stored at -80°C for later use. The CTAB method was used to extract the DNA of the elm ear. The extraction effect was detected by agarose gel electrophoresis, and the DNA mass concentration was determined by ultraviolet spectrophotometer. The DNA concentration was diluted to 100 ng / μL and placed at -80°C for later use.

[0031] (2) Screening and synthesis of SSR primers

[0032] Based on the SSR primers developed from the whole genome sequence of Auricularia auricula obtained earlier by the research team, three strains were used to screen SSR primers with clear banding, good amplification effect, repeatability and good polymorphism for subsequent experiments and analysis.

[0033] (3) PCR amplification and product detection

[0034] The PCR reaction system was: 5 μL of 10 nmol / μL dNTP Mixture, 0.8 μL of 10 pmol / μL SSR upstream and downstream primers, 1 μL of 100 ng / μL DNA, and 3.4 μL of dd H2O, for a total of 11 μL.

[0035] The PCR reaction conditions were as follows: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 45 s, annealing at 60°C for 45 s, extension at 72°C for 1 min, for a total of 35 cycles; extension at 72°C for 10 min; and storage at 4°C.

[0036] The detection method of polyacrylamide gel electrophoresis was referenced, and the silver staining method was referenced to Sanguinetti CJ et al.

[0037] (4) Data processing and analysis

[0038] Band statistics: Directly count the band types based on the electrophoresis results. (1) Count the clearly identifiable bands. Do not count the unclear bands that cannot be accurately identified. (2) At the same position of mobility, if the width of one band is more than twice the width of another band, they are considered different bands. (3) Statistics use 0 / 1 binary data. The presence of a band at the same position is assigned a value of 1, and the absence of a band is assigned a value of 0. Construct a 0 / 1 matrix in an Excel spreadsheet with the statistical results.

[0039] Data processing: Dataformater was used to convert the binary 0 / 1 data into the required data format. POPGEN 32 software was used to calculate the number of alleles (Na), effective number of alleles (Ne), Shannon's information index (I), and allele frequency. PowerMarker V3.25 software was used to calculate the polymorphism information content (PIC) of each pair of SSR primers and the Nei's genetic distance (GD) between the test strains. The genetic similarity coefficient between the strains was calculated based on the Nei's genetic distance, and the UPGMA cluster diagram of the test strains was constructed.

[0040] (5) Construction of DNA fingerprint

[0041] After amplifying 100 ulm ear germplasms with SSR primers, the band size data was read, polymorphic fragments were screened, and the polymorphic fragments were converted into 0, 1 matrices in binary mode and recorded in an EXCEL table. The core primers were screened according to the results of SSR polymorphism analysis by combining primers and genotypes, and the polymorphic fragments amplified by the core primers were used to generate a digital fingerprint map in the form of 0 / 1 binary data, that is, to construct a molecular ID card for each strain. Finally, the fingerprint code and strain information were combined to generate the corresponding QR code fingerprint pattern online.

[0042] 3. Results and analysis

[0043] (1) Primer screening results

[0044] Among the SSR primers developed based on the whole genome sequence of Ulmus auricula obtained by the research team in the early stage, a total of 18 pairs of SSR primers with good polymorphism and 11 pairs of SSR primers distributed on 5 genetic linkage groups of Ulmus auricula and associated with agronomic traits were screened out, totaling 29 pairs, which were synthesized by Jilin Kumei Biotechnology Co., Ltd. The primer information is shown in Table 2.

[0045] Table 2 SSR primer information

[0046]

[0047]

[0048] (2) Screening of core primers

[0049] According to the site information results of 29 pairs of SSR primers amplification, the PIC value of each primer, the number of amplified alleles and other factors were comprehensively analyzed, and the primer combinations were used to analyze 100 strains of Ulmus pumila strains. The principle of distinguishing the most strains with the least primer combination was followed until all 100 strains were completely separated.

[0050] As shown in Table 3, 7 pairs of SSR primers can distinguish 73 strains, 8 pairs of primers can distinguish 85 strains, 9 pairs of primers can distinguish 95 strains, and 10 pairs of SSR primers can completely distinguish 100 strains of Ulmus auricula. The polymorphic information content is 0.7185-0.8909, the primer with the largest PIC value is SSR558, and the primer SSR147 has the smallest PIC value. These 10 pairs of SSR primers can be used as core primers to completely distinguish 100 test strains and construct a two-dimensional code fingerprint.

[0051] The bands amplified by the 10 pairs of SSR primers in 100 strains of Ulmus auricula were recorded, and the digital DNA fingerprint information of 100 copies of Ulmus auricula was collected as the basis for the molecular ID string. Figure 4 As shown in Table 4, taking the amplification product of primer SSR558 as an example, the digital DNA fingerprint information of each strain in the region of about 60 to 100 bp is read, that is, the molecular ID string. The digital DNA fingerprint information of each strain is shown in Table 4. Figure 5 As shown, taking the amplification product of primer SSR500 as an example, the digital DNA fingerprint information of each strain in the region of about 60 to 90 bp, that is, the molecular ID string, is read. The digital DNA fingerprint information of each strain is shown in Table 4.

[0052] Table 3 SSR primer combinations for distinguishing 100 Ulmus auricula germplasm resources

[0053] Primer combinations Differentiate the number of strains SSR383 11 SSR383+SSR558 20 SSR383+SSR558+SSR572 29 SSR383+SSR558+SSR572+SSR197 38 SSR383+SSR558+SSR572+SSR197+SSR500 48 SSR383+SSR558+SSR572+SSR197+SSR500+SSR451 59 SSR383+SSR558+SSR572+SSR197+SSR500+SSR451+SSR397 73 SSR383+SSR558+SSR572+SSR197+SSR500+SSR451+SSR397+SSR147 85 SSR383+SSR558+SSR572+SSR197+SSR500+SSR451+SSR397+SSR147+SSR351 95 SSR383+SSR558+SSR572+SSR197+SSR500+SSR451+SSR397+SSR147+SSR351+SSR326 100

[0054] (3) Construction of DNA fingerprint

[0055] The selected core primers were used to generate a molecular ID card in 0-1 binary format, which is the SSR fingerprint code (Table 4). Finally, each germplasm fingerprint code was entered into the online website (http: / / cli.im / ) to generate a specific QR code ( Figure 3 ). The QR code information includes strain number, strain type, product characteristics information and fingerprint code, etc.

[0056] Table 4 Digital fingerprints of 100 ulmaria germplasm resources

[0057]

[0058]

[0059]

[0060]

[0061] (4) SSR primer polymorphism analysis

[0062] As shown in Table 5, 29 pairs of SSR primers detected a total of 272 allele loci in 100 tested materials. The number of allele loci amplified by each pair of primers ranged from 4 to 13. On average, each pair of primers detected 9.3793 allele variation loci, with abundant polymorphic loci and a percentage of 100%. The number of effective alleles (Ne) ranged from 2.6828 to 10.0857, with the largest number of alleles in primer SSR326 at 10.0857 and the smallest number in primer SSR1 at 2.6828, with an average value of 6.8574. Shanoon's information index (I) ranged from 1.1121 to 2.3842, with the highest number in SSR397 and the lowest in SSR1, with an average value of 1.9926. The observed heterozygosity (Ho) was 0. The expected heterozygosity (He) ranged from 0.6304 to 0.9054, with an average of 0.8412. The primer SSR326 had the highest value of 0.9054. The polymorphism information content (PIC) of primer allele sites varied from 0.5581 to 0.8922, with an average of 0.8161. The primer SSR225 had the highest value of 0.8922, and the primer SSR1 had the lowest value of 0.5581. The SSR polymorphism information content was not consistent with the allele variation. In general, the 29 pairs of SSR primers screened in this study amplified relatively rich polymorphisms and could be used for the study of genetic diversity of the test materials.

[0063] Table 5 SSR primer polymorphism information

[0064]

[0065] The change of PIC value is closely related to the number of alleles, but the change trend of the two is not positively correlated (Table 5). Although the number of alleles of SSR383, SSR451, SSR558 and SSR572 is large, their PIC values ​​are not the largest, while the PIC value of primer SSR225 with a medium number of alleles is the largest. Figure 1 Statistical analysis of the allele frequencies of 29 primer pairs revealed that the allele frequency distribution of most primers was uneven, especially in primers with a small number of alleles. For example, the frequency of allele B of SSR1 was as high as 48.5%, while the frequency of allele A was only 4%.

[0066] (5) Cluster analysis based on SSR molecular markers

[0067] The similarity coefficient can directly reflect the genetic differences between strains. The smaller the genetic distance, the closer the relationship between the strains; vice versa. Based on the genotype data of 29 pairs of polymorphic SSR markers amplified on the test materials, the Nei's genetic distance was calculated using Powermarker software and a cluster diagram of the test strains was constructed. The UPGMA cluster analysis results showed that ( Figure 2 ), the genetic distance between the 100 strains of Ulmus auricula was between 0.0039 and 0.4232, and the genetic differences between the strains were relatively obvious. At the genetic distance of 0.0039, the 100 strains of Ulmus auricula can be divided into three categories, among which the first group is backcross strains, the second group is parent strains and some backcross strains, and the third group is self-cross strains and a few backcross strains.

[0068] Group I includes 54 backcross strains, but from the clustering results, they are not clearly divided into two categories according to the different recurrent parents. It is preliminarily speculated that this may be due to the crossover of the genetic background of the non-recurrent parents of these strains. Group II includes C, D, 126-15 and 7 backcross strains. C and D are the recurrent parents of the backcross strains, and 126-15 is the non-recurrent parent. The backcross strains are all obtained by backcrossing the recurrent parent C and 126-15. The higher the similarity, the smaller the genetic distance between the varieties. It is inferred that the genetic background of these 7 strains is extremely similar to that of C, so they are clustered into one category. Cluster III included 33 self-fertilized strains and three backcross strains, GH89, GH112, and GH113, and was divided into two subgroups with a threshold of 0.0154, of which GH89 was clustered into one group alone and the other subgroup included 35 strains, including two backcross strains: GH112 and GH113 and GZ102 were derived from the same parent, Q186, so the genetic distance between these three strains was relatively close. Overall, the genetic background similarity of these strains varied, depending on which parent strains they were associated with.

[0069] It should be noted that when the claims of the present invention involve numerical ranges, it should be understood that the two endpoints of each numerical range and any numerical value between the two endpoints can be selected. In order to avoid redundancy, the present invention describes a preferred embodiment.

[0070] Although the preferred embodiments of the present invention have been described, those skilled in the art may make other changes and modifications to these embodiments once they have learned the basic creative concept. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.

[0071] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. A combination of SSR marker primers for ulmus aurantium germplasm resources, characterized in that: The SSR marker primer set includes primer set 1 whose sequences are shown in SEQ ID NO.1-2, primer set 2 as shown in SEQ ID NO.3-4, primer set 3 as shown in SEQ ID NO.5-6, primer set 4 as shown in SEQ ID NO.7-8, primer set 5 as shown in SEQ ID NO.9-10, primer set 6 as shown in SEQ ID NO.11-12, primer set 7 as shown in SEQ ID NO.13-14, primer set 8 as shown in SEQ ID NO.15-16, primer set 9 as shown in SEQ ID NO.17-18, and primer set 10 as shown in SEQ ID NO.19-20.

2. A kit, characterized in that: The kit comprises the SSR marker primer combination of ulmus fuciformis germplasm resources described in claim 1.

3. Use of the SSR marker primer combination of ulmus fusilli germplasm resources described in claim 1 or the kit described in claim 2 in constructing a fingerprint map of ulmus fusilli germplasm resources.

4. Use of the SSR marker primer combination of ulmus fusilli germplasm resources described in claim 1 or the kit described in claim 2 in identifying ulmus fusilli germplasm resources.

5. A method for constructing a fingerprint of ulmus auricula germplasm resources, characterized in that: The following steps are involved: The DNA of the elm ear germplasm resources is amplified by PCR using the SSR marker primer combination of the elm ear germplasm resources described in claim 1, the band size data is read, the polymorphic fragments are screened and obtained, and the polymorphic fragments are converted into 0, 1 matrix data in a binary manner to obtain the molecular ID fingerprint of the elm ear germplasm resources.

6. The method for constructing the fingerprint of the ulmus auricula germplasm resources according to claim 5, characterized in that: The reaction system of the PCR amplification is 5 μL of dNTP Mixture, 0.8±0.1 μL of forward primer, 0.8±0.1 μL of reverse primer, 1±0.1 μL of DNA, and ultrapure water added to 11 μL.

7. The method for constructing the fingerprint of ulmus auricula germplasm resources according to claim 5, characterized in that: The reaction conditions of the PCR amplification were as follows: pre-denaturation at 94° C. for 5 min; denaturation at 94° C. for 45 s, annealing at 60° C. for 45 s, extension at 72° C. for 1 min, for a total of 35 cycles; and extension at 72° C. for 10 min.

8. Application of the construction method described in claim 5, 6 or 7 in identifying Auricularia auricularia germplasm resources.

9. A method for identifying ulmus auricula germplasm resources, characterized in that: The following steps are involved: The SSR marker primer combination of the elm ear germplasm resources described in claim 1 is used to perform PCR amplification on the DNA of the elm ear germplasm resources to be tested, and the obtained PCR amplification band is compared with the elm ear germplasm resources fingerprint map constructed by the construction method described in claim 5, 6 or 7 to identify the type of the elm ear germplasm resources.

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