A Tool for the Design of the Minimal Fingerprinting SNP Set: Use Case for Barley
Abstract
:1. Introduction
2. Materials and Methods
2.1. The MDSearch Algorithm
2.2. Testing Dataset
2.3. Illumina Reads Preparation and Mapping
2.4. SNP-Calling and Filtration for Minimal Discriminatory SNP Set Selection
3. Results and Discussion
3.1. Filtration and Identification of Highly Polymorphic SNPs
- Initially, from a set of 850,174 detected mutations, 148,108 indels and complex mutations were removed (--snps-only), retaining only SNPs.
- Subsequently, out of 702,066 SNPs, 7252 multiallelic SNPs were removed (--min-alleles 2 --max-alleles 2), eliminating SNPs that discriminated more than two alleles.
- The SNPs were then sorted based on their ability to discriminate between all 254 studied varieties (--geno 0), resulting in the removal of SNPs that were unable to distinguish between all cultivars (SNP call rate < 100%). After this step, 41,112 SNPs remained.
- The remaining variants were filtered by minor allele frequency (MAF; --maf 0.01). Minor allele frequency indicates the prevalence of the reference variant of SNP. For highly polymorphic single-nucleotide mutations without homozygous genotypes in diploid species, the MAF cannot exceed 0.5, meaning that 50% of varieties have a reference SNP variant, and 50% have an alternative one. After appropriate evaluation, 37,450 SNPs with MAF < 1% were removed because such a low polymorphism frequency may indicate a genotyping error. Before the next stage, the set included 3662 SNPs.
- Homozygous SNPs are the most suitable for creating the SNP barcode, so 3166 SNPs that were heterozygous in at least one of the cultivars were removed from the set (--keep <filename>). After this procedure, 496 homozygous SNPs remained in the set.
- For certification, it is important to use SNPs that are not inherited linked, because this may affect the accuracy of the passport during the variety cultivation. Therefore, the pairwise linkage of 496 SNPs was analyzed in a sliding window of 50 SNPs with a step of 5 SNPs (--indep-pairwise 50 5 0.2). As a result, SNP pairs with linkage greater than 0.2 were removed, meaning that the genetic proximity of the two genotypes was at least 0.2. After removing 319 SNPs, 177 SNPs remained for further analysis.
- The optimal set of markers is considered to be one in which the markers are evenly distributed throughout the genome, which also reduces the risk of having linked markers in the set. Therefore, at the final stage of filtering, SNPs with a distance of less than 10 Mbp (--bp-space 10,000,000) were removed. As a result, 89 SNPs were excluded from the analysis, and the remaining 88 SNPs were utilized to identify the minimum discriminatory set of SNPs.
3.2. A Minimal SNP Set Development
3.3. Developed SNP Set Comparison with Known Sets
4. Conclusions
Author Contributions
Funding
Data Availability Statement
Conflicts of Interest
References
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# SNP | Chromosome Coordinate a | Ref. Allele | Alt. Allele | MAF b | PIC c |
---|---|---|---|---|---|
1 | 1H:5,550,434 | T | A | 0.11 | 0.20 |
2 | 1H:402,453,297 | G | A | 0.36 | 0.46 |
3 | 2H:39,017,947 | T | C | 0.11 | 0.20 |
4 | 2H:81,911,835 | G | T | 0.19 | 0.31 |
5 | 2H:560,080,229 | A | G | 0.38 | 0.47 |
6 | 3H:205,461,025 | C | G | 0.24 | 0.36 |
7 | 3H:447,564,794 | C | A | 0.23 | 0.35 |
8 | 3H:525,419,066 | G | C | 0.38 | 0.47 |
9 | 3H:566,880,335 | T | C | 0.27 | 0.39 |
10 | 3H:615,767,065 | T | C | 0.28 | 0.40 |
11 | 4H:453,072,008 | T | C | 0.15 | 0.26 |
12 | 5H:456,058,599 | G | C | 0.43 | 0.49 |
13 | 5H:519,706,238 | A | G | 0.13 | 0.23 |
14 | 5H:545,182,553 | T | C | 0.11 | 0.20 |
15 | 6H:13,191,080 | T | G | 0.44 | 0.49 |
16 | 6H:71,795,477 | A | T | 0.45 | 0.50 |
17 | 7H:208,590,826 | A | G | 0.12 | 0.21 |
18 | 7H:457,261,245 | G | C | 0.38 | 0.47 |
19 | 7H:616,514,313 | G | A | 0.38 | 0.47 |
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Ermolaev, A.; Samarina, M.; Strembovskiy, I.; Kroupin, P.; Karlov, G.; Kharchenko, P.; Voronov, S.; Eroshenko, L.; Kryuchenko, E.; Laptina, Y.; et al. A Tool for the Design of the Minimal Fingerprinting SNP Set: Use Case for Barley. Agronomy 2024, 14, 1802. https://doi.org/10.3390/agronomy14081802
Ermolaev A, Samarina M, Strembovskiy I, Kroupin P, Karlov G, Kharchenko P, Voronov S, Eroshenko L, Kryuchenko E, Laptina Y, et al. A Tool for the Design of the Minimal Fingerprinting SNP Set: Use Case for Barley. Agronomy. 2024; 14(8):1802. https://doi.org/10.3390/agronomy14081802
Chicago/Turabian StyleErmolaev, Aleksey, Mariya Samarina, Ilya Strembovskiy, Pavel Kroupin, Gennady Karlov, Pyotr Kharchenko, Sergey Voronov, Lyubov Eroshenko, Elizaveta Kryuchenko, Yulia Laptina, and et al. 2024. "A Tool for the Design of the Minimal Fingerprinting SNP Set: Use Case for Barley" Agronomy 14, no. 8: 1802. https://doi.org/10.3390/agronomy14081802