New Insights into the Genome Organization of Yeast Killer Viruses Based on “Atypical” Killer Strains Characterized by High-Throughput Sequencing
Abstract
:1. Introduction
2. Results
2.1. Phenotypic and Genotypic Characterization of S. cerevisiae and T. delbrueckii Killer Yeasts
2.2. Analysis of the dsRNA Sequence from ScV-M and TdV-M Viruses
2.3. Analysis of 5′- and 3′-Extra Sequences of M-Genomes
2.4. Analysis of Common Core Sequence of M-Genomes
2.5. Analysis of ScV-M and TdV-M Preprotoxin ORF Sequences
3. Discussion
3.1. Phenotypic and Genotypic Characterization of Atypical Killer Yeasts
3.2. Analysis of dsRNA Sequences from M Viruses
3.3. Sequence Comparison of the Killer Preprotoxins
4. Conclusions
5. Materials and Methods
5.1. Yeast Strains and Media
5.2. Determination of Yeast Killer Activity
5.3. Total Nucleic Acid Preparation and Nuclease Digestion
5.4. Nucleic Acid Analysis for Killer Yeast Typing
5.5. Viral dsRNA Purification
5.6. Preparation and Sequencing of cDNA Libraries from Purified Viral dsRNA
5.7. Viral dsRNA Sequence Assembly
5.8. Real-Time Quantitative PCR (qPCR) Conditions and Analysis
5.9. Miscellaneous
5.10. Nucleotide Sequence Accession Number
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Virus | Previous Estimated Size/Sequenced Length (bp)/Yeast Strain/ΔG | Newly Analysed Yeast Strain | Killer Phenotype/dsRNA Isotype | Size Estimated/Length (bp) Sequenced/ΔG | 5′-Extra Sequence (bp)/% Identity with: Size [Position] | 3′-Extra Sequence (bp)/% Identity with: Size [Position] | Stem-Loop Involving 5′-Extra Sequence [Position] (ΔG) | Stem-Loop Involving 3′-Extra Sequence [position] (ΔG) |
---|---|---|---|---|---|---|---|---|
M1 | 1830/1801/S.c. TF325/−1807 | S.c. EX231 | K1*/M1-1 | 1300/1933/−1937 | 66/94% S.c. LBC-2 virus, 66 nt [A(−)66 to T(−)1] | 66 /no identity found | Not found | Not found |
T.d. EX1257 | Kbarr-2/M1-2 | 1700/1933/−1916 | 66/no identity found | 65/94% S.c. 26S rRNA, 77 nt [C1783 to G(+)57] | Not found | Not found | ||
M2 | 1700/1163 + 209/S.c. 1384/−1861 | S.c. EX88 | K2/M2-3 | 1650/1625/−2020 | 33/no identity found | None | [A(−)23 to T126] (−69.5) | Not found |
S.c. EX1125 | K2/M2-4 | 1750/1723/−2297 | 87/no identity found | 44 /no identity found | [T(−)42 to A145] (−205) | [C1603 to G(+)39] (−137) | ||
Mlus | 2300/2033/S.c. EX229/−1970 | S.c. EX229 | Klus/Mlus-4 | 2300/2314/−2242 | 50/no identity found | 230/98% S.c. 16S mit rRNA, 242 nt [C2022 to G(+)203] | [C(−)50 to G107] (−30.1) | Not found |
S.c. EX436 | Klus/Mlus-1 | 2000/2346/−2409 | 51/93% Vitis vinifera, 47 nt [A(−)30 to T17], and 85% Saccharomycopsis fibuligera, 45 nt [A(−)30 to A15] | 198/100% S.c. 18S rRNA, 198 nt [A(+)1 to G(+)198] | [C(−)51 to G107] (−46.0) | Not found | ||
S.c. EX1160 | Klus*/Mlus-A | 2100/2268/−2723 | 218/no identity found | 17/no identity found | [T(−)125 to A97] (−246), or [A(−)67 to T129] (−62.8) | Not found | ||
S.c. EX1160 | Klus*/Mlus-B | 1700/1937/−2066 | 77/no identity found | None | Not found | Not found | ||
S.c. EX1160 | Klus*/Mlus-C | 1350/2055/−1962 | 22/no identity found | None | Not found | Not found | ||
Mbarr-1 | 1700/1705/T.d. EX1180/−1727 | T.d. EX1257 | Kbarr-2*/Mbarr-2B | 1300/1835/−1878 | 64/88% Cucumis melo, 41 nt [A(−)18 to G23] | 66/97% T.d. (95% S.c.) 26S rRNA, 76 nt [G1759 to C(+)65] | [T(−)2 to A53] (−40.6) | Not found |
Strain | Genotype [Relevant Phenotype] | Origin |
---|---|---|
Sc EX88 * | MAT a/α HO/HO cyhS/cyhS M2-3 [K2+] | M. Ramírez a (from wine) |
Sc EX85 * | MAT a/α HO/HO cyhS/cyhS LA M2-3 [K2+] | M. Ramírez a (from wine) |
Sc EX85R * | MAT a/α HO/HO CYHR/cyhS M20 [cyhR K20] | M. Ramírez a (from EX85) |
Sc EX229 * | MAT a/α HO/HO cyhS/cyhS LA LBC Mlus-4 [Klus+] | M. Ramírez a (from wine) |
Sc EX229-R1 * | MAT a/α HO/HO CYHR/cyhS [cyhR Klus0] | M. Ramírez a (from EX229) |
Sc EX33 * | MAT a/α HO/HO [LA0 K10 K20 K280 Klus0] | M. Ramírez a (from wine) |
Sc EX73 * | MAT a/α HO/HO LA M2-3 [K2+] | M. Ramírez a (from wine) |
Sc EX198 * | MAT a/α HO/HO LA LBC Mlus-3 [Klus+] | M. Ramírez a (from wine) |
Sc EX231 | MAT a/α HO/HO LA LBC M1-1 [K1+] | This study (from wine) |
Sc EX436 | MAT a/α HO/HO LA Mlus-1 [Klus+] | This study (from wine) |
Sc EX1125 | MAT a/α HO/HO LA LBC M2-4 [K2+] | This study (from wine) |
Sc EX1160 | MAT a/α HO/HO LA LBC Mlus-A Mlus-B Mlus-C[Klus+] | This study (from wine) |
Sc F166 * | MAT α leu1 kar1 LA-HNB M1 [K1+] | J.C. Ribas b (from R. Wickner) |
Sc F182 * | MAT α his2 ade1 leu2-2 ura3-52 ski2-2 LA M28 [K28+] | J. C. Ribas b (from M. Schmitt) |
Td EX1180 * | wt LAbarr-1 Mbarr-1 [Kbarr-1+] | M. Ramírez a (from wine) |
Td EX1180-11C4 * | cyhR LAbarr-1 Mbarr-1 [cyhR Kbarr-1+] | M. Ramírez a (from EX1180) |
Td EX1180-2K− * | cyhR LAbarr-1 Mbarr-10 [cyhR Kbarr0] | M. Ramírez a (from EX1180) |
Td EX1257 | wt LAbarr-2 M1-2 Mbarr-2B [Kbarr-2+] | This study (from wine) |
Td EX1257-CYH5 | cyhR LAbarr-2 M1-2 Mbarr-2B [cyhR Kbarr-2+] | M. Ramírez a (from EX1257) |
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Ramírez, M.; Velázquez, R.; López-Piñeiro, A.; Naranjo, B.; Roig, F.; Llorens, C. New Insights into the Genome Organization of Yeast Killer Viruses Based on “Atypical” Killer Strains Characterized by High-Throughput Sequencing. Toxins 2017, 9, 292. https://doi.org/10.3390/toxins9090292
Ramírez M, Velázquez R, López-Piñeiro A, Naranjo B, Roig F, Llorens C. New Insights into the Genome Organization of Yeast Killer Viruses Based on “Atypical” Killer Strains Characterized by High-Throughput Sequencing. Toxins. 2017; 9(9):292. https://doi.org/10.3390/toxins9090292
Chicago/Turabian StyleRamírez, Manuel, Rocío Velázquez, Antonio López-Piñeiro, Belén Naranjo, Francisco Roig, and Carlos Llorens. 2017. "New Insights into the Genome Organization of Yeast Killer Viruses Based on “Atypical” Killer Strains Characterized by High-Throughput Sequencing" Toxins 9, no. 9: 292. https://doi.org/10.3390/toxins9090292