Okay, let's tackle this question step by step. The user wants to know how the pathway enrichment analysis was conducted in the given scientific article. They provided a detailed Methods section, so I need to go through that carefully. First, question 1 asks what tool was used for pathway enrichment analysis. In the Methods section under "De novo assembly and functional annotation," there's a mention of KEGG pathway annotation using Blast all software against the KEGG database. So the tool here is KEGG, but the actual analysis might have been done using another tool. Wait, no. The sentence says they used Blast all software against the KEGG database for pathway annotation. So the tool for pathway enrichment would be the KEGG database itself, and the method is using Blast all. Hmm, but maybe the actual enrichment analysis is done with another tool. Let me check again. The text says: "KEGG pathways with a Q-value ≤0.05 are significantly enriched in DEGs." The Q-value usually refers to FDR, so the statistical test is part of the KEGG analysis. The tool might be the KEGG pathway annotation tool. But the user might be referring to the specific software used for the enrichment. Wait, the article says "KEGG pathway annotation using Blast all software against the KEGG database." So the tool is Blast all, and the gene set library is KEGG. So answer 1 is Blast all software. Question 2: Was a tool version number provided? The article mentions "Blast all software" but doesn't specify a version number. So the answer is no, version not provided. Question 3: What gene set library was queried? The answer is KEGG, as mentioned in the Methods section. Question 4: Was a background gene list defined? The article doesn't explicitly mention defining a background gene list. They talk about DEGs and pathways, but not a specific background. So answer is not described. Question 5: What statistical test was used? The text mentions KEGG pathways with a Q-value ≤0.05. Q-value is associated with false discovery rate correction. The statistical test is probably a hypergeometric test or similar, but the article doesn't specify the exact test. However, it does mention Q-value, which is related to FDR. But the question is about the statistical test. Since the article doesn't specify the test, the answer is not described. Question 6: Was FDR correction used? The article says "KEGG pathways with a Q-value ≤0.05 are significantly enriched." Q-value is a measure of FDR, so yes, FDR correction was used. Wait, let me verify each point again. For the tool, the article says "KEGG pathway annotation using Blast all software against the KEGG database." So the tool is Blast all. The gene set library is KEGG. For the statistical test, they mention Q-value, which is FDR. But the actual test isn't specified. So answer 5 is not described. For question 6, since Q-value is used, which is FDR, the answer is yes. So compiling the answers: 1. Tool: Blast all software 2. Version: Not provided 3. Gene set library: KEGG 4. Background gene list: Not described 5. Statistical test: Not described 6. FDR correction: Yes, via Q-value Here is the collated information based on the provided article: | **Question** | **Answer** | |--------------|------------| | 1. What tool was used for pathway enrichment analysis? | **Blast all software** was used for KEGG pathway annotation. | | 2. Was a tool version number provided? | **No**, the article does not specify a version number for the tool. | | 3. What gene set library was queried? | **KEGG (Kyoto Encyclopedia of Genes and Genomes)** database was queried. | | 4. Was a background gene list defined for pathway enrichment analysis? | **Not described**; the article does not explicitly mention defining a background gene list. | | 5. What statistical test was used for enrichment analysis? | **Not described**; the article does not specify the exact statistical test (e.g., hypergeometric test) used for enrichment. It only mentions that pathways were considered significant based on a **Q-value ≤ 0.05**. | | 6. Was false discovery rate correction used? | **Yes**, the article states that pathways with a **Q-value ≤ 0.05** were considered significantly enriched, which implies the use of false discovery rate (FDR) correction to control false positives. | ### Notes: - The **Q-value** mentioned in the article is a standard metric for FDR correction, confirming the use of FDR in pathway enrichment analysis. - The **Blast all software** was used to annotate pathways against the KEGG database, but no further details about the enrichment algorithm or test statistics are provided.