Abstract
Premise of the Study
Novel and cost‐effective microsatellite markers were developed to explore the population genetics, biogeographic structure, and evolutionary history of the prized Euro‐Asian wild edible ectomycorrhizal fungus Tricholoma matsutake (Tricholomataceae).
Methods and results
Eighteen new polymorphic simple sequence repeat loci, detected from a microsatellite‐enriched genomic library, were used to characterize 131 individuals from eight T. matsutake populations. The number of alleles ranged from two to 10, with averages of 1.42 to 3.22. Levels of observed and expected heterozygosity ranged from 0.00–1.00 and from 0.00–0.83, with mean values of 0.21 and 0.26, respectively. In total, 50% of the loci showed interspecific transferability and polymorphism in the related species T. equestre.
Conclusions
These newly developed markers will aid research into the genetic diversity and population structure of T. matsutake. They can also be used in other species of Tricholoma.
Keywords: capillary electrophoresis, population genetics, simple sequence repeat (SSR), Tricholoma matsutake, Tricholomataceae
The ectomycorrhizal basidiomycete Tricholoma matsutake (S. Ito & S. Imai) Sing (Tricholomataceae) is one of the most expensive edible mushrooms (Wang et al., 1997). No fruiting body of T. matsutake has ever been artificially produced to date. Price fluctuations and the question of how to ensure the sustainable use of the resource in the context of climate change and varying management approaches are of major concern. For many mountainous communities in the Himalayas, harvesting the matsutake mushroom is a major source of household income. This has led to intensive harvesting of this fungus in its natural habitat, which places its sustainability at risk. Therefore, to ensure better management of T. matsutake, a thorough understanding of its biology, ecology, and population genetics is essential.
Tricholoma matsutake has a Eurasian distribution (Matsushita et al., 2005) and is mainly found in eastern Asia, the Himalayan region, and northern Europe. Previous molecular studies have focused on developing markers to track the geographic origin of the matsutake mushroom (Xu and Hong, 2007) and to differentiate between populations at regional levels (Lian et al., 2003). The rapid development of whole genomic sequencing has afforded researchers greater opportunities to explore more novel and cost‐effective simple sequence repeat (SSR) markers. Moreover, capillary electrophoresis in combination with fluorescence‐labeled SSR markers has made it possible to identify complex aneupolyploid hybrids and carry out genetic evaluations (Pan et al., 2003). In this study, we attempt to detect reliable SSRs by analyzing the whole genomic sequencing data of T. matsutake and the related species T. equestre (L.) P. Kumm. The results of the study will enable us to understand the biogeographic patterns and evolutionary processes of T. matsutake, as well as facilitate its sustainable utilization.
METHODS AND RESULTS
Ten polymorphic SSR markers (Appendix S1) of T. matsutake have been previously reported (Lian et al., 2003). However, only 60% of these loci displayed polymorphism (Appendix S1) when tested against our 131 samples from Europe and Asia. We developed additional primers for T. matsutake using whole‐genome sequencing data (175.76 Mbp) of T. matsutake published by the Genome Portal of the Department of Energy's Joint Genome Institute (National Center for Biotechnology Information [NCBI] accession no. PRJNA200596) to construct an enriched microsatellite library. The MIcroSAtellite Identification Tool (MISA; Thiel et al., 2003) was employed to detect SSRs with the criteria of eight, five, five, five, and five repeat units for di‐, tri‐, tetra‐, penta‐, and hexanucleotide motifs, respectively. In total, 7983 SSRs were identified from whole genome sequence data, the most common being trinucleotide repeats (87.9%, 7025), followed by hexanucleotides (11.1%, 885), dinucleotides (0.5%, 42), tetranucleotides (0.3%, 22), and pentanucleotides (0.1%, 9). A total of 48 primer pairs were designed by Primer Premier 5 software (PREMIER Biosoft International, Palo Alto, California, USA), using the following parameters: primer length 18–21 bp with the amplified product size set to range from 100 to 700 bp. Of these primer pairs, 21 (18 polymorphic and three monomorphic; Table 1) amplified successfully (GenBank accession no. KY986283–KY986303), and the remaining 27 failed to amplify (Appendix S2). These 18 polymorphic primer pairs were tested for polymorphism using 131 individuals collected from eight distinct geographic locations in Bhutan, China, the Republic of Korea, Japan, and neighboring populations in Finland and Sweden (Appendix 1). Fresh matsutake mushrooms were collected in the field where possible. However, to maximize geographic representation, we also purchased mushrooms from different collectors at surrounding township‐level local markets.
Table 1.
Locus | Primer sequences (5′–3′) | Repeat motif | Allele size range (bp) | T a (°C) | GenBank accession no. |
---|---|---|---|---|---|
LJW075 | F: GGGAATGGAGATGCTGAG | (CAGGGA)6 | 387–405 | 55 | KY986294 |
R: TTGTTGTGATGAGGGTAGGA | |||||
LJW068 | F: GTGTCACCGCCGCCAGTAT | (TCCCAA)5 | 387–411 | 57 | KY986293 |
R: TCGGAGACGCTCGGATGCT | |||||
LJW045 | F: GGGAATGGAGATGCTGAGR: TTGTTGTGATGAGGGTAGGA | (TCCAGC)5 | 387–405 | 63 | KY986289 |
LJW089 | F: AGAGCGTCATTGCTTGGG | (GGGACT)6 | 186–210 | 55 | KY986298 |
R: CTGTCGGATGCCTCGTAG | |||||
LJW002 | F: AGCCAAACACCAAAGCCCAACA | (CCTCTC)5 | 215–233 | 56 | KY986283 |
R: CGCCCACAGCCGCATAAA | |||||
LJW014 | F: CCGTATTCTTCCTTTCGTTG | (CAC)10 | 162–192 | 54 | KY986285 |
R: CTGCCTTCTTACCGCCAC | |||||
LJW100 | F: CAAGTCCACCTCGTTTCTC | (CACAAC)6 | 284–338 | 59 | KY986299 |
R: AATATCCATAACTACGCCTGA | |||||
LJW018 | F: GCAGATTCGCACCAGGAT | (CCTCTC)5 | 303–327 | 50 | KY986286 |
R: CGCCCACAGCCGCATAAA | |||||
LJW036 | F: CTTGACGGAAGAAAGAGTATGT | (AGCAGG)6 | 315–339 | 54 | KY986287 |
R: CGTGAGCCGAGTGGTGAT | |||||
LJW052 | F: GTGTCACCGCCGCCAGTAT | (TCCCAA)5 | 387–411 | 56 | KY986290 |
R: TCGGAGACGCTCGGATGCT | |||||
LJW053 | F: GTGCGGAACCATCTCAGTC | (CAA)9 | 365–380 | 54 | KY986291 |
R: CGTAGGAGCGTCCATAGTGT | |||||
LJW077 | F: ACAACACCAATGCCAACC | (CACCAA)5 | 328–358 | 48 | KY986295 |
R: CAAGAAATGAGAAACAAAA | |||||
LJW079 | F: GTGTCACCGCCGCCAGTAT | (TCCCAA)5 | 387–411 | 57 | KY986296 |
R: TCGGAGACGCTCGGATGCT | |||||
LJW104 | F: CCACCTAACACCCACTCTT | (TCTCAC)5 | 291–309 | 50 | KY986300 |
R: GACAGCACGGAACCATCT | |||||
LJW005 | F: TTGGTGAAGGCGGGAAGA | (GAGAGG)6 | 142–154 | 59 | KY986284 |
R: CATGCCACTCATAGGCAGTA | |||||
LJW083 | F: TCATCGTTCAACTGTGGCTTCT | (ACC)5(CCT)5 | 201–213 | 50 | KY986297 |
R: CGTTTGTGGCGGCTATTT | |||||
LJW145 | F: CCCCTCCCAACTCAACAT | (GGTGTT)7 | 210–252 | 56 | KY986302 |
R: CGGCGTAACTGCACTAACAT | |||||
LJW154 | F: GCTTTGTCTCAGCCTTCAAAG | (GCTGGT)6 | 115–175 | 55 | KY986303 |
R: AAGACAACCACAAATCCTCCC | |||||
LJW39* | F: TCACTTTGGGAGTCTGTC | (AAAGGG)5 | 394 | 54 | MF318475 |
R: GTTTGCTTATTTGTTGGGTA | |||||
LJW76* | F: GAGGATTGCCTGAGTGAT | (GAAAGA)10 | 413 | 57 | MF318484 |
R: AACCTGCTTATGTGGATTTT | |||||
LJW41* | F: TCACTTTGGGAGTCTGTC | (AAAGGG)5 | 227 | 54 | MF318476 |
R: GTTTGCTTATTTGTTGGGTA |
T a = annealing temperature.
Values are based on 131 samples.
*Monomorphic loci.
Genomic DNA was extracted from silica‐dried cap tissue following the cetyltrimethylammonium bromide (CTAB) method of Doyle and Doyle (1987). In addition, individuals of T. equestre (n = 15, collected from specimens deposited at the herbarium of the Kunming Institute of Botany, Chinese Academy of Sciences [KUN]) were chosen to test the cross‐species amplification of polymorphic and monomorphic markers in T. matsutake. Voucher specimens were deposited at KUN (Appendix 1).
PCR amplification was performed in a 25‐μL reaction mixture that consisted of 12.5 μL of 2× Taq Master Mix (total 1 mL of solution containing 100 units Taq polymerase, 0.5 mM dNTP, 20 mM Tris‐HCl [pH 8.3], 3 mM MgCl2; Vazyme Biotech Co., Nanjing, China), 1 μL of each primer, and 1.5 μL of genomic DNA (~50 ng/μL). The 5′ end primers of SSRs (Table 1) were labeled with two different fluorescent dyes (6‐FAM and HEX) under the following conditions: 95°C for 3 min; 35 cycles of 95°C for 15 s, the appropriate annealing temperatures (Table 1) for 30 s, and 72°C for 16 s; and a final extension of 72°C for 5 min.
The product was analyzed by capillary electrophoresis on an ABI 3730xl DNA analyzer (Applied Biosystems, Foster City, California, USA) at Beijing Microread Genetech Co. (Beijing, China) using GeneScan 500 ROX Size Standard (Applied Biosystems). SSR fragment lengths were analyzed by GeneMapper version 3.2 (Applied Biosystems). Aberrant peaks were not scored (Pan et al., 2003). We calculated the number of alleles, as well as observed (H o) and expected heterozygosities (H e) for each population. Hardy–Weinberg equilibrium was determined using GenAlEx version 6.4 (Peakall and Smouse, 2012).
Of the 48 primer pairs, 18 (37.5%; Table 1) were polymorphic when screened using 131 individuals from eight populations. The number of alleles ranged from two to 10 and averages ranged from 1.42 to 3.22. Levels of H o and H e varied from 0.00 to 1.00 and 0.00 to 0.83, respectively (Table 2), with mean values of 0.21 to 0.26. Eight out of the 18 polymorphic SSR loci showed significant deviations from Hardy–Weinberg equilibrium in different populations (P < 0.05) (Table 2). Within T. equestre, 50% of the SSR primer pairs were successfully cross‐amplified, with levels of H o and H e ranging from 0.06–0.93 and 0.32–0.83 (Table 2).
Table 2.
T. matsutake | |||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Locus | BTN‐TP (n = 15) | CN‐TB (n = 14) | CN‐SC (n = 15) | CN‐YN (n = 15) | CN‐NC (n = 15) | KOR‐SE (n = 14) | JPN‐NK (n = 13) | FIN‐RM (n = 15) | T. equestre (n = 15) | ||||||||||||||||||
A | H o | H e | A | H o | H e | A | H o | H e | A | H o | H e | A | H o | H e | A | H o | H e | A | H o | H e | A | H o | H e | A | H o | H e | |
LJW75 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.07 | 0.06 | 1 | 0.00 | 0.00 | 3 | 0.60 | 0.58** | 1 | 0.00 | 0.00** | 4 | 0.38 | 0.65** | 0 | 0.00 | 0.00** | 0 | 0.00 | 0.00** |
LJW68 | 2 | 0.08 | 0.07 | 1 | 0.00 | 0.00 | 2 | 0.20 | 0.28 | 2 | 0.27 | 0.32 | 4 | 0.20 | 0.70*** | 3 | 0.25 | 0.32* | 4 | 0.67 | 0.69 | 2 | 0.20 | 0.18 | 0 | 0.00 | 0.00 |
LJW45 | 2 | 0.07 | 0.06 | 2 | 0.10 | 0.10 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 3 | 0.33 | 0.62** | 3 | 0.42 | 0.34 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 0 | 0.00 | 0.00 |
LJW89 | 5 | 0.20 | 0.30 | 4 | 0.38 | 0.56*** | 4 | 0.53 | 0.50*** | 5 | 0.67 | 0.56 | 3 | 0.50 | 0.54 | 4 | 0.50 | 0.62 | 3 | 0.50 | 0.41 | 1 | 0.00 | 0.00 | 0 | 0.00 | 0.00 |
LJW2 | 2 | 0.00 | 0.49*** | 2 | 0.00 | 0.17*** | 2 | 0.00 | 0.12*** | 2 | 0.00 | 0.12*** | 3 | 0.58 | 0.47 | 1 | 0.00 | 0.00 | 3 | 0.33 | 0.61 | 2 | 0.57 | 0.49 | 0 | 0.00 | 0.00 |
LJW14 | 3 | 0.09 | 0.31*** | 2 | 0.08 | 0.20** | 2 | 0.07 | 0.06 | 4 | 0.13 | 0.29*** | 3 | 0.71 | 0.53 | 5 | 0.90 | 0.69 | 3 | 0.00 | 0.65*** | 4 | 0.18 | 0.38*** | 0 | 0.00 | 0.00*** |
LJW100 | 3 | 0.21 | 0.31 | 4 | 0.30 | 0.47*** | 4 | 0.15 | 0.21*** | 4 | 0.20 | 0.30** | 5 | 0.29 | 0.41 | 4 | 0.14 | 0.26** | 3 | 0.67 | 0.61 | 2 | 0.14 | 0.13 | 0 | 0.00 | 0.00 |
LJW18 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 4 | 0.53 | 0.54 | 3 | 0.50 | 0.46 | 3 | 0.46 | 0.41 | 2 | 0.13 | 0.44*** | 7 | 0.54 | 0.45 |
LJW36 | 1 | 0.00 | 0.00 | 2 | 0.09 | 0.09 | 3 | 0.43 | 0.54 | 3 | 0.27 | 0.53 | 3 | 0.53 | 0.64 | 3 | 0.79 | 0.63 | 5 | 0.38 | 0.71** | 1 | 0.00 | 0.00** | 5 | 0.87 | 0.68 |
LJW52 | 1 | 0.00 | 0.00 | 3 | 0.29 | 0.25 | 3 | 0.40 | 0.57 | 4 | 0.47 | 0.56 | 4 | 0.27 | 0.66*** | 3 | 0.57 | 0.52 | 5 | 0.69 | 0.76 | 1 | 0.00 | 0.00 | 6 | 0.29 | 0.78 |
LJW53 | 2 | 0.00 | 0.48*** | 2 | 0.07 | 0.07 | 2 | 0.20 | 0.18 | 2 | 0.07 | 0.28*** | 4 | 0.33 | 0.29 | 3 | 0.36 | 0.30 | 3 | 0.40 | 0.46 | 1 | 0.00 | 0.00 | 2 | 0.18 | 0.46** |
LJW77 | 4 | 0.31 | 0.28 | 1 | 0.00 | 0.00 | 3 | 0.36 | 0.39 | 4 | 0.33 | 0.39** | 4 | 0.27 | 0.34** | 3 | 0.36 | 0.39 | 3 | 0.17 | 0.52** | 1 | 0.00 | 0.00** | 0 | 0.00 | 0.00** |
LJW79 | 3 | 0.29 | 0.41 | 3 | 0.29 | 0.64** | 4 | 0.40 | 0.68*** | 6 | 0.53 | 0.66 | 6 | 0.27 | 0.72** | 4 | 0.43 | 0.64* | 6 | 0.62 | 0.75 | 4 | 0.20 | 0.51*** | 6 | 0.11 | 0.72*** |
LJW104 | 2 | 0.60 | 0.46 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.13 | 0.23 | 3 | 0.67 | 0.54 | 3 | 0.86 | 0.64 | 3 | 0.31 | 0.32 | 1 | 0.00 | 0.00 | 3 | 0.38 | 0.32 |
LJW5 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.08 | 0.07 | 1 | 0.00 | 0.00 | 2 | 0.08 | 0.41*** |
LJW83 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.13 | 0.12 | 2 | 0.00 | 0.12*** | 1 | 0.00 | 0.00*** | 2 | 0.00 | 0.46*** | 1 | 0.00 | 0.00*** | 10 | 0.13 | 0.83*** |
LJW145 | 2 | 0.14 | 0.13 | 3 | 0.29 | 0.45** | 4 | 0.27 | 0.34** | 3 | 0.47 | 0.38 | 3 | 0.53 | 0.42 | 3 | 0.43 | 0.36 | 3 | 0.38 | 0.32 | 1 | 0.00 | 0.00 | 3 | 0.00 | 0.59*** |
LJW154 | 3 | 0.13 | 0.13 | 3 | 0.07 | 0.14*** | 5 | 0.33 | 0.35*** | 4 | 0.13 | 0.24** | 4 | 0.14 | 0.20*** | 3 | 0.14 | 0.40** | 2 | 0.00 | 0.43*** | 1 | 0.00 | 0.00*** | 0 | 0.00 | 0.00*** |
Trma01 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.27 | 0.23 | 2 | 0.08 | 0.07 | — | — | — | 2 | 0.00 | 0.12** | 2 | 0.00 | 0.12** |
Trma02 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.87 | 0.49*** | 2 | 1.00 | 0.50*** | 2 | 0.38 | 0.31 | — | — | — | 2 | 0.00 | 0.24*** | 0 | 0.00 | 0.00 |
Trma06 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.07 | 0.36*** | 2 | 0.15 | 0.43** | 0 | 0.00 | 0.00 | — | — | — | 0 | 0.00 | 0.00 | 0 | 0.00 | 0.00 |
Trma07 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.07 | 0.36*** | 2 | 0.08 | 0.45*** | 0 | 0.00 | 0.00 | — | — | — | 0 | 0.00 | 0.00 | 0 | 0.00 | 0.00 |
Trma08 | 2 | 0.00 | 0.14** | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 3 | 0.00 | 0.34*** | 1 | 0.00 | 0.00 | 0 | 0.00 | 0.00 | — | — | — | 0 | 0.00 | 0.00 | 0 | 0.00 | 0.00 |
Trma10 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.07 | 0.07 | — | — | — | 1 | 0.00 | 0.00 | 0 | 0.00 | 0.00 |
Trma12 | 2 | 0.79 | 0.50 | 2 | 0.57 | 0.46 | 2 | 0.36 | 0.38 | 2 | 0.33 | 0.28 | 2 | 0.73 | 0.49 | 2 | 0.17 | 0.15 | — | — | — | 2 | 0.13 | 0.12 | 1 | 0.00 | 0.00 |
Trma13 | 1 | 0.00 | 0.00 | 0 | 0.00 | 0.00 | 1 | 0.00 | 0.00 | 2 | 0.33 | 0.28 | 2 | 0.20 | 0.28 | 2 | 0.25 | 0.22 | — | — | — | 2 | 0.38 | 0.31 | 0 | 0.00 | 0.00 |
Trma14 | 2 | 0.00 | 0.12** | 1 | 0.00 | 0.00 | 2 | 0.00 | 0.13** | 3 | 0.73 | 0.66 | 2 | 0.60 | 0.46 | 3 | 0.36 | 0.45 | — | — | — | 2 | 0.67 | 0.44 | 0 | 0.00 | 0.00 |
Trma16 | 2 | 0.93 | 0.50*** | 2 | 0.64 | 0.48 | 2 | 0.93 | 0.50*** | 2 | 1.00 | 0.50*** | 2 | 0.93 | 0.50*** | 2 | 1.00 | 0.50*** | — | — | — | 2 | 1.00 | 0.50*** | 2 | 1.00 | 0.50*** |
— = monomorphic locus; A = number of alleles sampled; H e = expected heterozygosity within populations; H o = observed heterozygosity.
Locality and voucher information are provided in Appendix 1.
Significant deviation from Hardy–Weinberg equilibrium (*P < 0.1, **P < 0.05, ***P < 0.01).
Conclusions
The 18 newly reported polymorphic SSR markers for T. matsutake are reliable and will be used in the further study of the species. The interspecific transferability and polymorphism shown in the related species T. equestre suggest that these markers may also be applicable to the study of genetic diversity in other Tricholoma species.
Data accessibility
Sequence information for the developed primers has been deposited to the National Center for Biotechnology Information (NCBI); GenBank accession numbers are provided in Table 1.
Supporting information
Acknowledgments
This project was supported by the National Natural Science Foundation of China (grant no. 31370513), the Chinese Academy of Sciences “Light of West China” Program, and the Southeast Asia Biodiversity Research Institute, Chinese Academy of Sciences (2015CASEABRIRG001). The authors thank Dr. Zhikun Wu and Jinchao Yang (Kunming Institute of Botany, Chinese Academy of Sciences), Niclas Bergius (County Administrative Board of Västmanland, Sweden), and Prof. Chunlan Lian (University of Tokyo) for their kind support in providing the field collection samples, as well as Dr. Andrew W. Stevenson for assistance with English editing.
Appendix 1. Collection information for all Tricholoma samples used in this study.
Species | Population code | Collection locationa | Geographic coordinates | Voucher specimen accession no. | N |
---|---|---|---|---|---|
Tricholoma matsutake (S. Ito & S. Imai) Sing | BTN‐TP | Paro dzongkhag, Kingdom of Bhutan | 27°33′47″N, 90°53′46″E | X. F. Yang et al. 100087 | 15 |
CN‐TB | Nyingchi, Tibet Autonomous Region, People's Republic of China | 29°56′38″N, 94°47′56″E | X. F. Yang et al. 100118 | 14 | |
CN‐SC | Yajiang, Sichuan Province, People's Republic of China | 30°01′52″N, 101°00′50″E | X. F. Yang et al. 100192 | 15 | |
CN‐YN | Deqing, Yunnan Province, People's Republic of China | 28°29′10″N, 98°54′40″E | X. F. Yang et al. 100219 | 15 | |
CN‐NC | Antu, Jilin Province, People's Republic of China | 43°06′47″N, 128°53′53″E | X. F. Yang et al. 100453 | 15 | |
KOR‐SE | Cheongju Province, Republic of Korea | 36°49′59″N, 127°45′18″E | X. F. Yang et al. 100479 | 14 | |
JPN‐NK | Nagano, Japan | 43°28′44″N, 129°19′48″E | — | 13 | |
FIN‐RM | Kalix Municipality, Norrbotten County, Sweden | 65°51′00″N, 23°10′12″E | — | 5 | |
Rovaniemi Municipality, Finland | 66°30′00″N, 25°43′59″E | — | 5 | ||
Nuuksio National Park, Espoo, Finland | 60°18′0.13″N, 24°27′59″E | — | 5 | ||
Tricholoma equestre (L.) P. Kumm. | CN‐YNE | Kunming, Yunnan Province, People's Republic of China | 25°9′47″N, 102°43′48″E | Y. Q. Xiao et al. 49796 | 15 |
The DNA samples from Japan and Sweden were contributed by Dr. Chunlan Lian and Niclas Bergius, respectively.
Li, J.‐W. , Yin X., Zhao Y.‐J., Yang S.‐J., Vaario L.‐M., and Yang X.‐F.. 2018. Microsatellite markers for the prized matsutake mushroom (Tricholoma matsutake, Tricholomataceae). Applications in Plant Sciences 6(12): e1202.
These authors contributed equally to this work and should be considered co‐first authors.
Notes
N = number of individuals.
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Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Supplementary Materials
Data Availability Statement
Sequence information for the developed primers has been deposited to the National Center for Biotechnology Information (NCBI); GenBank accession numbers are provided in Table 1.