Transcriptome Sequencing on Six Agaricus bisporus Strains at Four Developmental Stages
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摘要:
目的 通过多个双孢蘑菇菌株不同发育阶段的差异转录组分析，为进一步验证双孢蘑菇发育相关基因及探讨其分子机理奠定基础。 方法 对双孢蘑菇主栽品种As2796及其亲本02、8213，其回交子代W192，以及国外野生菌株ARP159、国内野生菌株AgLH830共6个具有重要代表性的菌株子实体原基期、幼菇期、采摘期、开伞期等4个不同发育阶段共24个样品进行转录组测序，并与双孢蘑菇参考基因组序列进行比对，根据比对结果进行各基因在不同样品中的表达量分析及差异表达基因识别，发掘新基因与共同基因的差异表达，并进行各数据库的基因功能注释。 结果 结果共鉴定到10 660个转录本，发掘新基因677个，其中237个得到功能注释。与原基期相比，6个菌株在幼菇期、采摘期和开伞期分别有49、82、73个共同差异表达基因，其中有13个是相同的基因。 结论 发现了一批在双孢蘑菇子实体不同发育阶段具有显著差异表达的基因，筛选出不同菌株不同阶段的共有差异基因，对双孢蘑菇子实体发育中重要的差异基因进行了注释与探讨。 Abstract:Objective Transcriptome analysis was conducted on Agaricus bisporus at 4 developmental stages to identify the genes associated with and decipher the molecular mechanism involving the fungal development. Method From 6 representative strains of A. bisporus including main cultivar As2796, its parents 02 and 8213, its backcrossing offspring W192, foreign wild strain ARP159, and domestic wild strain AgLH830, at primordium, young, harvesting, and opening stages, 24 fruiting body specimens were collected for transcriptome sequence analysis. By aligning them against the reference genome sequence of A. bisporus, the genes that were differentially expressed were identified. Both unique and common differentially expressed genes (DEGs) were clearly exposed to be annotated using the databases to determine their specific functions. Result Among the 10 660 transcripts obtained, 677 genes were unique with 237 functionally annotated. The tested A. bisporus shared 49 common DEGs between primordium and young stages, 82 between primordium and harvest stages, and 73 between primordium and pileus opening stages. And, 13 genes were found commonly present in the various strains. Conclusion Both unique and common DEGs in A. bisporus at the 4 developmental stages were identified and annotated in the study. -
图 1 双孢蘑菇6个菌株原基期与幼菇期共同差异表达基因的COG分类
Figure 1. COG annotation of common DEGs between primordium and young stages in 6 strains of A. bisporus
图 2 双孢蘑菇6个菌株原基期与幼菇期共同差异表达基因的GO富集分析
Figure 2. GO enrichment analysis of common DEGs between primordium and young stages in 6 strains of A. bisporus
图 3 双孢蘑菇6个菌株原基期与幼菇期共同差异表达基因的KEGG通路类型
Figure 3. KEGG pathway of common DEGs between primordium and young stages in 6 strains of A. bisporus
表 1 样品测序数据及与参考基因组的序列比对结果统计
Table 1. Statistics of experimental data and alignment with reference on genome sequence
编号
ID样品
Samples总片读数
Total reads匹配片读数
Mapped reads匹配读段
Mapped reads/%≥Q30的百分比
≥Q30 Percenlagel/%GC含量
GC Content/%T01 2796原基 35666572 26745594 74.99 87.10 49.06 T02 2796幼菇 35313022 27062540 76.64 86.34 49.39 T03 2796采收 38280048 29421017 76.86 86.59 49.27 T04 2796开伞 38250636 28047724 73.33 86.57 49.55 T05 02原基 33427134 28017783 83.82 86.06 49.26 T06 02幼菇 35819542 30247728 84.44 87.01 49.44 T07 02采收 34688248 29636455 85.44 88.16 49.49 T08 02开伞 31922230 26803817 83.97 86.88 49.51 T09 192原基 34556016 27403923 79.30 89.03 49.45 T10 192幼菇 33710834 26856893 79.67 88.61 49.41 T11 192采收 33484752 26652110 79.59 88.04 49.63 T12 192开伞 35109700 27405076 78.06 87.62 49.59 T13 8213原基 62520314 44230857 70.75 91.10 49.87 T14 8213幼菇 37020010 28479519 76.93 87.00 49.36 T15 8213采收 48831096 37862207 77.54 91.00 49.92 T16 8213开伞 33504694 25964053 77.49 89.57 49.49 T17 AgLH830原基 46629820 33805235 72.50 91.37 49.90 T18 AgLH830幼菇 37064508 25651715 69.21 90.87 50.06 T19 AgLH830采收 60230312 43341259 71.96 92.94 49.64 T20 AgLH830开伞 59003490 42371223 71.81 93.21 49.65 T21 ARP159原基 58535934 43943896 75.07 93.01 49.41 T22 ARP159幼菇 57898228 42151349 72.80 92.16 49.80 T23 ARP159采收 50652184 37439256 73.91 91.69 49.66 T24 ARP159开伞 52116880 37313086 71.60 90.30 49.72 
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表 2 新基因功能注释结果统计
Table 2. Number of new genes with functional annotation
注释数据库
Annotated databases新基因数目
New gene numberGO 14 COG 10 KEGG 25 Swiss-Prot 24 NR 236 All 237
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表 3 差异表达基因数目统计
Table 3. Number of DEGs
差异表达基因集
DEG Set差异表达基因总数
All DEGs上调基因数目
up-regulated DEGs下调基因数目
down-regulated DEGsT01_vs_T02 632 168 464 T01_vs_T03 790 241 549 T01_vs_T04 843 378 465 T05_vs_T06 866 499 367 T05_vs_T07 951 467 484 T05_vs_T08 892 501 391 T09_vs_T10 665 197 468 T09_vs_T11 784 297 487 T09_vs_T12 864 432 432 T13_vs_T14 724 325 399 T13_vs_T15 816 357 459 T13_vs_T16 904 403 501 T17_vs_T18 597 238 359 T17_vs_T19 1012 486 526 T17_vs_T20 806 310 496 T21_vs_T22 673 227 446 T21_vs_T23 813 302 511 T21_vs_T24 870 499 371
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表 4 注释的差异表达基因数量统计
Table 4. Number of annotated DEGs
差异表达基因集
DEG Set获注释DEG
Annotated DEG注释数据库Annotated databases GO COG KEGG Swiss-Prot NR T01_vs_T02 607 214 249 157 307 607 T01_vs_T03 764 263 301 201 376 764 T01_vs_T04 823 324 348 262 430 823 T05_vs_T06 841 361 378 360 502 841 T05_vs_T07 908 358 389 334 503 908 T05_vs_T08 839 320 331 267 419 839 T09_vs_T10 649 254 270 193 353 649 T09_vs_T11 759 283 320 215 397 759 T09_vs_T12 844 316 348 237 422 844 T13_vs_T14 712 313 328 309 424 712 T13_vs_T15 797 324 350 287 438 797 T13_vs_T16 870 323 364 266 447 870 T17_vs_T18 581 222 256 160 302 581 T17_vs_T19 973 315 360 250 440 972 T17_vs_T20 781 325 349 250 421 781 T21_vs_T22 658 292 318 234 395 658 T21_vs_T23 796 342 376 271 466 796 T21_vs_T24 854 363 387 298 487 854
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表 5 双孢蘑菇子实体发育阶段共同差异基因及在菌株As2796中的表达量
Table 5. Thirteen common DEGs among 6 strains of A. bisporus at 4 developmental stages and their expression levels in As2796
基因编号
Gene ID相对表达量FPKM 基因长度
Length/bp基因注释
AnnotationT01 T02 T03 T04 estExt_fgenesh2_kg.C_50657 2800.3 363.3 111.4 199.6 2497 para-aminobenzoate synthetase e_gw1.3.1175.1 189.9 16.7 8.2 9.3 1459 Null estExt_Genewise1.C_180108 644.4 31.9 13.2 43.8 1014 sterigmatocystin biosynthesis
peroxidase stcC (Precursor)fgenesh2_pm.2_863 469.9 74.9 39.3 48.1 4105 Long-chain-fatty-acid-CoA ligase estExt_fgenesh2_pg.C_20058 172.0 11.6 31.4 5.4 1224 Null estExt_Genewise1.C_51672 301.1 24.0 8.4 32.8 2719 transcriptional enhancer factor estExt_fgenesh2_kg.C_50125 191.4 34.0 12.7 10.4 3434 Null e_gw1.1.1627.1 84.8 18.7 16.0 13.2 557 Null estExt_Genewise1Plus.C_160094 38.8 5.4 4.4 3.3 2329 O-methylsterigmatocystin oxidoreductase estExt_Genewise1Plus.C_60597 492.6 50.8 34.4 87.3 6696 Zinc/cadmium resistance protein estExt_fgenesh2_pg.C_180054 41.7 1.8 0.2 4.3 453 Null estExt_fgenesh2_pm.C_170010 28.2 1.6 0.7 0.3 2198 Uncharacterized transporter estExt_fgenesh2_kg.C_10275 117.6 955.7 1528.6 1515.5 2478 aminodeoxychorismate synthase
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