| 草菇磷酸果糖激酶（PFK）基因克隆、结构及不同菌株中表达量分析 |
| Cloning, structural analyses and expression levels of phosphofructokinase gene in different strains of Volvariella volvacea |
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| 中文关键词:草菇，基因结构，可变剪切，协同增效，异核体 |
| 英文关键词:Volvariella volvacea, gene model, alternative splicing, synergistic gene expression effect, heterokaryon |
| 基金项目:国家现代农业产业技术体系建设专项资金（No. CARS 24） |
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| 中文摘要: |
| 从福建主栽品种屏优一号PY分离到2个不能正常出菇的单孢菌株PYd21、PYd15，二者配对杂交得到可正常出菇异核菌株H15-21。用solexa测序技术对PYd21基因组从头测序（de novo sequencing），对PYd15、PYd21、H15-21的菌丝体分别进行数字基因表达谱测序，对8个样品（PYd15、PYd21、H15-21的菌丝体，H15-21出菇后的原基、钮扣期菌柄、蛋形期菌柄、伸长期菌柄和成熟期菌柄）mRNA等量混合物进行转录组测序。克隆测序了PYd21与PYd15中磷酸果糖激酶（PFK）基因，比对结果表明二者序列一致。利用转录组数据对PFK基因结构进行分析，结果表明：该基因全长3,494bp，有12个外显子、11个内含子；开放阅读框（ORF）全长2,457bp，编码818个氨基酸，5′端非翻译区（5′UTR）长度281bp，3′端非翻译区（3′UTR）全长103bp；RNA在加工过程中存在1种可变剪切类型、6个可变剪切位点。数字基因表达谱分析结果表明，PFK基因在PYd21、PYd15及H15-21中的标准表达量分别为71.08、120.61、251.85（transcript per million clean tags，TPM），表达量依次升高，与菌丝生长速度显著正相关。并且异核菌株H15-21表达量高于2个单孢菌株之和，基因表达存在协同增效作用。采用实时荧光定量PCR技术对基因表达量进行验证，表达谱数据切实可靠。 |
| 英文摘要: |
| Two single spore unfruitful isolates, PYd21 and PYd15, were obtained from Volvariella volvacea CV. PY in Fujian Province. A hybrid heterokaryon, H15-21, was obtained by mating PYd21 and PYd15, and it could fruit normally. Using solexa genome analyzer platform, the genome of PYd21 was de novo sequencing; digital gene expression profiles of the mycelium including PYd21, PYd15 and H15-21 were sequenced respectively; a transcriptome containing equivalent mRNA of 8 samples including the mycelium samples of PYd21, PYd15 and H15-21, fruiting body samples of “pinhead” stage, and stipes of “button”, “egg”, “elongation” and “mature” stages from H15-21 were sequenced. Phosphofructokinase (PFK) genes were cloned and sequenced from both PYd21 and PYd15, and their structures were analyzed through paired-end mapping of transcriptional reads. Sequence alignment revealed both sequences to be identical, and PFK consists of 3,494bp with an open reading frame (ORF) of 2,457bp, encoding a polypeptide of 818 amino acids. PFK contained 12 exons and 11 introns, and the 5′UTR and 3′UTR were 281bp and 103bp long, respectively. One type of alternative splicing and 6 alternative splicing sites were identified during RNA processing. TPM (transcripts per million tags) values for PYd21, PYd15 and H15-21 were 71.08, 120.61 and 251.85, respectively, showing significant positive correlation to the growth rate of mycelium. The data indicated that the expression level of PFK gene in H15-21 had synergistic effect. The expression levels of PFK gene were confirmed by real-time quantitative PCR. |
| 刘朋虎,谢宝贵,邓优锦,江玉姬. 草菇磷酸果糖激酶（PFK）基因克隆、结构及不同菌株中表达量分析[J]. 菌物学报, 2013, 32(2): 253-260 |
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