Add Vercel configuration for rewrites and build settings

2025-12-17 17:47:45 +08:00 · 2025-12-17 17:47:45 +08:00 · 0d31d42de4
parent 072524e303
commit 0d31d42de4
32 changed files with 7435 additions and 289 deletions
--- a/.gitignore
+++ b/.gitignore
@ -6,8 +6,9 @@
 01.reference/*
 02.mapping_assembly/*
 03.denovo_assembly/*
-04.plastid/*
+04.cds_prediction/*
 05.reduce_redundancy/*
+05.orthology_inference/*
 06.phylogeny_reconstruction/*
 98.results/*
 99.scripts/bucky/
@ -15,3 +16,4 @@
 .pueue.yml
 run.status
 !*/_description.md
+tmp/
--- a/98.results/_description.md
+++ b/98.results/_description.md
@ -0,0 +1,5 @@
+# Results Directory
+This directory contains the results generated from the data analysis, which should be uploaded to the public sharing platform.
+
+## Files Included
+
--- a/99.scripts/miscs/hmmsearch_result_to_new_ogs_dir.py
+++ b/99.scripts/miscs/hmmsearch_result_to_new_ogs_dir.py
@ -0,0 +1,316 @@
+#! /usr/bin/env python3
+import os
+import re
+import argparse
+import sys
+from Bio import SeqIO
+from Bio.SeqRecord import SeqRecord
+from Bio.Seq import Seq
+from pathlib import Path
+import logging
+
+
+logging.basicConfig(level=logging.DEBUG, format="[%(levelname)s]: %(message)s")
+
+
+class BestHit:
+    def __init__(self, e_value: float | None = None, target_name: str | None = None):
+        self.e_value = e_value
+        self.target_name = target_name
+
+
+def parse_fasta(fasta_file_path) -> dict[str, str]:
+    """
+    Parse a FASTA file and return a list of sequence ids.
+    Args:
+        fasta_file_path: Path to the FASTA file
+    Returns:
+        list: Dictionary of sequences
+    """
+    sequences: dict[str, str] = {}
+    try:
+        for record in SeqIO.parse(fasta_file_path, "fasta"):
+            sequences[record.id] = str(record.seq)
+    except Exception as e:
+        logging.error(f"Error parsing FASTA file {fasta_file_path}: {e}")
+    return sequences
+
+
+def get_ids_from_fasta(fasta_file_path) -> list[str]:
+    """
+    Parse a FASTA file and return a list of sequence ids.
+    Args:
+        fasta_file_path: Path to the FASTA file
+    Returns:
+        list: List of sequence ids
+    """
+    seqs = parse_fasta(fasta_file_path)
+    return list(seqs.keys())
+
+
+def parse_hmmer_tbl(tbl_file_path) -> BestHit:
+    """
+    Parse HMMER tbl format result file and extract best hit information
+
+    Args:
+        tbl_file_path: Path to the tbl file
+
+    Returns:
+        dict: Best hit information
+    """
+    best_hit = BestHit()
+
+    try:
+        with open(tbl_file_path, "r") as f:
+            for line in f:
+                # Skip comment lines
+                if line.startswith("#"):
+                    continue
+
+                # Split line (tbl format is typically space or tab separated)
+                parts = re.split(r"\s+", line.strip())
+                if len(parts) < 5:
+                    continue
+
+                # Extract key information: target name, E-value, score, etc.
+                # tbl format columns: target name, target accession, query name, query accession, E-value, score, etc.
+                target_name = parts[0]
+                e_value = float(parts[4])  # Full sequence E-value
+
+                # If it's a new sequence or we found a better hit (lower E-value)
+                if best_hit.e_value is None or e_value < best_hit.e_value:
+                    best_hit = BestHit(e_value=e_value, target_name=target_name)
+    except Exception as e:
+        logging.error(f"Error parsing tbl file {tbl_file_path}: {e}")
+        return BestHit()
+
+    return best_hit
+
+
+def find_corresponding_files(fasta_dir, tbl_dir) -> list[tuple[Path, Path]]:
+    """
+    Find corresponding FASTA and tbl file pairs
+
+    Args:
+        fasta_dir: Directory containing FASTA files
+        tbl_dir: Directory containing tbl result files
+
+    Returns:
+        list: List of (fasta_file_path, tbl_file_path) tuples
+    """
+    file_pairs: list[tuple[Path, Path]] = []
+
+    # Get all FASTA files
+    fasta_files = {}
+
+    for fasta_path in Path(fasta_dir).glob("*.fa"):
+        stem = fasta_path.stem
+        fasta_files[stem] = fasta_path
+
+    # Find corresponding tbl files
+    for stem, fasta_path in fasta_files.items():
+        tbl_name = f"{stem}.fa.hmmsearch.tblout"
+        tbl_path = Path(tbl_dir) / tbl_name
+        if tbl_path.exists():
+            file_pairs.append((fasta_path, tbl_path))
+        else:
+            logging.warning(f"No corresponding tbl file found for {stem}")
+
+    return file_pairs
+
+
+def add_best_hit_to_ogs_dir(
+    file_pair: tuple[Path, Path],
+    all_sequences: dict[str, str],
+    ogs_dir: Path,
+    species_name: str | None = None,
+) -> tuple[str, list[str]]:
+    """
+    Add best hit information to FASTA file
+
+    Args:
+        file_pair: Tuple of (fasta_path, tbl_path)
+        all_sequences: Dictionary of homolog sequences
+        ogs_dir: Directory to save FASTA files
+        species_name: Species name of candidate species
+    Returns:
+        tuple: (stem, seq_ids)
+    """
+    fasta_path, tbl_path = file_pair
+
+    # Parse tbl file to get best hits
+    best_hit = parse_hmmer_tbl(tbl_path)
+
+    if not best_hit.target_name:
+        logging.warning(f"No valid hits found in {tbl_path}, skipping...")
+        return fasta_path.stem, []
+
+    best_seq = best_hit.target_name
+    seq_ids = get_ids_from_fasta(fasta_path)
+    seqs: dict[str, str] = {}
+    for seq_id in seq_ids:
+        seq = all_sequences.get(seq_id)
+        if not seq:
+            logging.warning(f"Sequence ID {seq_id} not found! Skipping...")
+            return fasta_path.stem, []
+        seqs[seq_id] = seq
+    modified_ogs_seq_record = [
+        SeqRecord(seq=Seq(v), id=k.split("@")[0], description="")
+        for k, v in seqs.items()
+    ]
+    homolog_dir = Path(ogs_dir) / fasta_path.stem
+    homolog_dir.mkdir(parents=True, exist_ok=True)
+    modified_ogs_seq_path = homolog_dir / f"{fasta_path.stem}.fa"
+    SeqIO.write(modified_ogs_seq_record, modified_ogs_seq_path, "fasta")
+
+    homolog_seq = all_sequences.get(best_seq)
+    if not homolog_seq:
+        logging.warning(f"Best hit sequence {best_seq} not found! Skipping...")
+        return fasta_path.stem, []
+
+    if not species_name:
+        species_name = best_seq.split("@")[0]
+    homolog_seq_path = homolog_dir / f"{fasta_path.stem}_{species_name}.fa"
+    homolog_seq_record = SeqRecord(
+        seq=Seq(homolog_seq), id=best_seq.split("@")[0], description=""
+    )
+    SeqIO.write([homolog_seq_record], homolog_seq_path, "fasta")
+    seq_ids.append(best_seq)
+    return fasta_path.stem, seq_ids
+
+
+def process_all_files(fasta_dir, tbl_dir, output_dir, all_fasta, species_name):
+    """
+    Process all FASTA and tbl file pairs
+
+    Args:
+        fasta_dir: Directory containing FASTA files
+        tbl_dir: Directory containing tbl result files
+        output_file: Output file path
+        all_fasta: FASTA format cds file of candidate species
+    """
+    logging.info("Starting file processing...")
+    logging.info(f"FASTA directory: {fasta_dir}")
+    logging.info(f"tbl directory: {tbl_dir}")
+    logging.info(f"Output Directory: {output_dir}")
+    logging.info(f"Homolog FASTA file: {all_fasta}")
+
+    og_list = {}
+    all_sequences = parse_fasta(all_fasta)
+    if not all_sequences:
+        logging.error(f"No sequences found in FASTA file {all_fasta}")
+        sys.exit(1)
+
+    # Find corresponding file pairs
+    file_pairs = find_corresponding_files(fasta_dir, tbl_dir)
+
+    if not file_pairs:
+        logging.error("No corresponding FASTA-tbl file pairs found")
+        sys.exit(1)
+
+    logging.info(f"Found {len(file_pairs)} file pairs to process")
+
+    output_file = Path(output_dir) / "updated_ogs_list.txt"
+    ogs_dir = Path(output_dir) / "ogs"
+    ogs_dir.mkdir(parents=True, exist_ok=True)
+
+    # Process each file pair
+    for i, file_pair in enumerate(file_pairs, 1):
+        logging.info(f"Processing pair {i}/{len(file_pairs)}:")
+        stem, seq_ids = add_best_hit_to_ogs_dir(
+            file_pair, all_sequences, ogs_dir, species_name
+        )
+        if not seq_ids:
+            logging.info(f"Skipping {stem}!")
+            continue
+        og_list[stem] = seq_ids
+
+    with open(output_file, "w") as out_f:
+        for og, ids in og_list.items():
+            out_f.write(f"{og}\t" + "\t".join(ids) + "\n")
+
+    logging.info(f"\nProcessing completed! All results saved to: {output_dir}")
+    logging.info(f"Updated OGS list saved to: {output_file}")
+    logging.info(f"Total OGS processed successfully: {len(og_list)}")
+
+
+def main(fasta_directory, tbl_directory, output_directory, all_fasta, species_name):
+    """
+    Main function - take paths and run processing
+
+    Args:
+        fasta_directory: Folder containing FASTA files
+        tbl_directory: Folder containing tbl result files
+        output_file: Output file path
+        all_fasta: FASTA format cds file of candidate species
+        species_name: Species name of candidate species
+    """
+    # Check if input directories exist
+    if not os.path.exists(fasta_directory):
+        logging.error(f"FASTA directory does not exist {fasta_directory}")
+        sys.exit(1)
+
+    if not os.path.exists(tbl_directory):
+        logging.error(f"tbl directory does not exist {tbl_directory}")
+        sys.exit(1)
+
+    try:
+        Path(output_directory).mkdir(parents=True, exist_ok=True)
+    except Exception as e:
+        logging.error(f"Error creating output directory {output_directory}: {e}")
+        sys.exit(1)
+
+    if not os.path.exists(all_fasta):
+        logging.error(f"Homolog FASTA file does not exist {all_fasta}")
+        sys.exit(1)
+
+    # Process all files
+    process_all_files(
+        fasta_directory, tbl_directory, output_directory, all_fasta, species_name
+    )
+
+
+if __name__ == "__main__":
+    # Parse command-line arguments and call main
+    parser = argparse.ArgumentParser(
+        description="Add HMMER best hit into orthologs FASTA."
+    )
+    parser.add_argument(
+        "-d",
+        "--fasta_directory",
+        required=True,
+        help="Directory containing FASTA files",
+    )
+    parser.add_argument(
+        "-t",
+        "--tbl_directory",
+        required=True,
+        help="Directory containing tbl result files",
+    )
+    parser.add_argument(
+        "-f",
+        "--all_fasta",
+        required=True,
+        help="FASTA format cds file of cadidate species",
+    )
+    parser.add_argument(
+        "-o",
+        "--output_directory",
+        required=True,
+        help="Directory to write output seqname",
+    )
+    parser.add_argument(
+        "-s",
+        "--species_name",
+        required=False,
+        help="Species name of candidate species",
+    )
+    args = parser.parse_args()
+
+    main(
+        args.fasta_directory,
+        args.tbl_directory,
+        args.output_directory,
+        args.all_fasta,
+        args.species_name,
+    )
--- a/99.scripts/miscs/hmmsearch_result_to_new_ogs_list.py
+++ b/99.scripts/miscs/hmmsearch_result_to_new_ogs_list.py
@ -1,215 +0,0 @@
-#! /usr/bin/env python3
-import os
-import re
-import argparse
-from Bio import SeqIO
-from pathlib import Path
-
-
-def parse_fasta(fasta_file_path):
-    """
-    Parse a FASTA file and return a list of sequence ids.
-    Args:
-        fasta_file_path: Path to the FASTA file
-    Returns:
-        list: List of sequence ids
-    """
-    sequence_ids = []
-    try:
-        for record in SeqIO.parse(fasta_file_path, "fasta"):
-            sequence_ids.append(record.id)
-    except Exception as e:
-        print(f"Error parsing FASTA file {fasta_file_path}: {e}")
-    return sequence_ids
-
-
-def parse_hmmer_tbl(tbl_file_path):
-    """
-    Parse HMMER tbl format result file and extract best hit information
-
-    Args:
-        tbl_file_path: Path to the tbl file
-
-    Returns:
-        dict: Best hit information
-    """
-    best_hit = {}
-
-    try:
-        with open(tbl_file_path, "r") as f:
-            for line in f:
-                # Skip comment lines
-                if line.startswith("#"):
-                    continue
-
-                # Split line (tbl format is typically space or tab separated)
-                parts = re.split(r"\s+", line.strip())
-                if len(parts) < 5:
-                    continue
-
-                # Extract key information: target name, E-value, score, etc.
-                # tbl format columns: target name, target accession, query name, query accession, E-value, score, etc.
-                target_name = parts[0]
-                e_value = float(parts[4])  # Full sequence E-value
-
-                # If it's a new sequence or we found a better hit (lower E-value)
-                if not best_hit.keys() or e_value < best_hit["e_value"]:
-                    best_hit = {
-                        "e_value": e_value,
-                        "target_name": target_name,
-                    }
-
-    except Exception as e:
-        print(f"Error parsing tbl file {tbl_file_path}: {e}")
-        return {}
-
-    return best_hit
-
-
-def find_corresponding_files(fasta_dir, tbl_dir):
-    """
-    Find corresponding FASTA and tbl file pairs
-
-    Args:
-        fasta_dir: Directory containing FASTA files
-        tbl_dir: Directory containing tbl result files
-
-    Returns:
-        list: List of (fasta_file_path, tbl_file_path) tuples
-    """
-    file_pairs = []
-
-    # Get all FASTA files
-    fasta_files = {}
-
-    for fasta_path in Path(fasta_dir).glob("*.fa"):
-        stem = fasta_path.stem
-        fasta_files[stem] = fasta_path
-
-    # Find corresponding tbl files
-    for stem, fasta_path in fasta_files.items():
-        tbl_name = f"{stem}.fa.hmmsearch.tblout"
-        tbl_path = Path(tbl_dir) / tbl_name
-        if tbl_path.exists():
-            file_pairs.append((fasta_path, tbl_path))
-        else:
-            print(f"Warning: No corresponding tbl file found for {stem}")
-
-    return file_pairs
-
-
-def add_best_hit_to_og_list(file_pair):
-    """
-    Add best hit information to FASTA file
-
-    Args:
-        file_pair: Tuple of (fasta_path, tbl_path)
-    Returns:
-        tuple: (stem, seq_ids)
-    """
-    fasta_path, tbl_path = file_pair
-
-    # Parse tbl file to get best hits
-    best_hit = parse_hmmer_tbl(tbl_path)
-
-    if not best_hit:
-        print(f"Warning: No valid hits found in {tbl_path}")
-        return fasta_path.stem, []
-
-    best_seq = best_hit["target_name"]
-    seq_ids = parse_fasta(fasta_path)
-    seq_ids.append(best_seq)
-
-    return fasta_path.stem, seq_ids
-
-
-def process_all_files(fasta_dir, tbl_dir, output_file):
-    """
-    Process all FASTA and tbl file pairs
-
-    Args:
-        fasta_dir: Directory containing FASTA files
-        tbl_dir: Directory containing tbl result files
-        output_file: Output file path
-    """
-    print("Starting file processing...")
-    print(f"FASTA directory: {fasta_dir}")
-    print(f"tbl directory: {tbl_dir}")
-    print(f"Output file: {output_file}")
-    print("-" * 50)
-
-    og_list = {}
-
-    # Find corresponding file pairs
-    file_pairs = find_corresponding_files(fasta_dir, tbl_dir)
-
-    if not file_pairs:
-        print("No corresponding FASTA-tbl file pairs found")
-        return
-
-    print(f"Found {len(file_pairs)} file pairs to process")
-
-    # Process each file pair
-    for i, file_pair in enumerate(file_pairs, 1):
-        print(f"\nProcessing pair {i}/{len(file_pairs)}:")
-        stem, seq_ids = add_best_hit_to_og_list(file_pair)
-        if not seq_ids:
-            print(f"Skipping {stem} due to no valid hits")
-            continue
-        og_list[stem] = seq_ids
-
-    with open(output_file, "w") as out_f:
-        for og, ids in og_list.items():
-            out_f.write(f"{og}\t" + "\t".join(ids) + "\n")
-
-    print(f"\nProcessing completed! All results saved to: {output_file}")
-
-
-def main(fasta_directory, tbl_directory, output_file):
-    """
-    Main function - take paths and run processing
-
-    Args:
-        fasta_directory: Folder containing FASTA files
-        tbl_directory: Folder containing tbl result files
-        output_file: Output file path
-    """
-    # Check if input directories exist
-    if not os.path.exists(fasta_directory):
-        print(f"Error: FASTA directory does not exist {fasta_directory}")
-        return
-
-    if not os.path.exists(tbl_directory):
-        print(f"Error: tbl directory does not exist {tbl_directory}")
-        return
-
-    # Process all files
-    process_all_files(fasta_directory, tbl_directory, output_file)
-
-
-if __name__ == "__main__":
-    # Parse command-line arguments and call main
-    parser = argparse.ArgumentParser(
-        description="Add HMMER best hit into orthologs FASTA."
-    )
-    parser.add_argument(
-        "-f",
-        "--fasta_directory",
-        required=True,
-        help="Directory containing FASTA files",
-    )
-    parser.add_argument(
-        "-t",
-        "--tbl_directory",
-        required=True,
-        help="Directory containing tbl result files",
-    )
-    parser.add_argument(
-        "-o",
-        "--output_file",
-        required=True,
-        help="File to write output seqname",
-    )
-    args = parser.parse_args()
-
-    main(args.fasta_directory, args.tbl_directory, args.output_file)
--- a/99.scripts/miscs/macse.sh
+++ b/99.scripts/miscs/macse.sh
@ -0,0 +1,28 @@
+#! /bin/bash
+
+FS_LR=7
+
+if [ "$#" -ne 3 ]; then
+    echo "Usage: $0 <reliable_seq> <less_reliable_seq> <stem>"
+    echo "Perform MACSE alignment on given sequences"
+    exit 1
+fi
+
+seq=$1
+seq_lr=$2
+stem=$3
+
+{
+    echo "Command:"
+    echo "macse -prog alignSequences -seq $seq -seq_lr ${seq_lr}"
+    echo "      -out_NT ${stem}.nal -out_AA ${stem}.pal"
+    echo "      -optim 2 -max_refine_iter 3 -local_realign_init 0.2 -fs_lr $FS_LR"
+} >alignSequences.log
+macse -prog alignSequences \
+    -seq "$seq" -seq_lr "${seq_lr}" \
+    -out_NT "$stem".nal -out_AA "$stem".pal \
+    -optim 2 \
+    -max_refine_iter 3 \
+    -local_realign_init 0.2 \
+    -fs_lr $FS_LR \
+    >>alignSequences.log 2>&1
--- a/99.scripts/miscs/rename_trinity_fasta.py
+++ b/99.scripts/miscs/rename_trinity_fasta.py
@ -7,6 +7,7 @@ Function: Rename sequences in FASTA file to format: [prefix@sequence_number]
 import sys
 import os

+
 def rename_fasta_sequences(input_file, prefix, output_file=None):
    """
    Rename sequence headers in a FASTA file
@ -22,25 +23,36 @@ def rename_fasta_sequences(input_file, prefix, output_file=None):
        file_base, file_ext = os.path.splitext(input_file)
        output_file = f"{file_base}_renamed{file_ext}"

+    match_tsv = f"{output_file}.tsv"
+    print(f"Input file: {input_file}")
+    print(f"Output file: {output_file}")
+    print(f"Naming format: {prefix}@mrna_<number>")
+    print(f"Match TSV file: {match_tsv}")
+
    # Counter for sequences
    seq_count = 0

    try:
-        with open(input_file, 'r') as fin, open(output_file, 'w') as fout:
+        with (
+            open(input_file, "r") as fin,
+            open(output_file, "w") as fout,
+            open(match_tsv, "w") as tsvout,
+        ):
+            tsvout.write("Original_Name\tNew_Name\n")
            for line in fin:
-                if line.startswith('>'):
+                if line.startswith(">"):
                    # Sequence header line: rename it
                    seq_count += 1
-                    new_name = f">{prefix}@mrna_{seq_count}\n"
-                    fout.write(new_name)
+                    original_name = line[1:].strip().split()[0]
+                    new_name = f"{prefix}@mrna_{seq_count}\n"
+                    fout.write(f">{new_name}")
+                    tsvout.write(f"{original_name}\t{new_name}\n")
                else:
                    # Sequence data line: write as-is
                    fout.write(line)

        print(f"Successfully renamed {seq_count} sequences")
-        print(f"Input file: {input_file}")
-        print(f"Output file: {output_file}")
-        print(f"Naming format: {prefix}@mrna_number")
+

    except FileNotFoundError:
        print(f"Error: Input file '{input_file}' not found")
@ -49,6 +61,7 @@ def rename_fasta_sequences(input_file, prefix, output_file=None):
        print(f"Error processing file: {e}")
        sys.exit(1)

+
 def main():
    """Main function"""
    if len(sys.argv) < 3:
@ -67,5 +80,6 @@ def main():

    rename_fasta_sequences(input_file, prefix, output_file)

+
 if __name__ == "__main__":
    main()
--- a/99.scripts/workflow/orthology_inference/01.hmmer_sc.sh
+++ b/99.scripts/workflow/orthology_inference/01.hmmer_sc.sh
@ -0,0 +1,51 @@
+#! /bin/bash
+set -e
+
+if [ "$#" -ne 4 ]; then
+    echo "Usage: $0 <ogs_dir> <outdir> <proteome> <threads>"
+    echo "search homologous sequences in <proteome> using HMMs built from orthogroup alignments"
+    exit 1
+fi
+
+ogs_dir=$(readlink -f "$1")
+outdir=$2
+proteome=$(readlink -f "$3")
+threads=$4
+
+mkdir -p "$outdir"
+cd "$outdir" || exit 1
+echo "Working directory: $(pwd)"
+echo "Using OGS directory: $ogs_dir"
+echo "Using $threads threads"
+echo ""
+echo "Starting orthogroup sequence alignment..."
+mkdir -p msa
+echo -n >mafft.cmds
+for i in "$ogs_dir"/*.fa; do
+    j=$(basename "$i")
+    echo "linsi --quiet $i > msa/$j" >>mafft.cmds
+done
+xargs -t -P "$threads" -I cmd -a mafft.cmds bash -c "cmd"
+echo "Orthogroup sequence alignment completed."
+echo ""
+echo "Starting HMM building from alignments..."
+mkdir -p hmms
+echo -n >hmmbuild.cmds
+for i in msa/*.fa; do
+    j=$(basename "$i")
+    echo "hmmbuild -o hmms/${j}.hmmbuild.out --amino hmms/${j}.hmm $i" >>hmmbuild.cmds
+done
+xargs -t -P "$threads" -I cmd -a hmmbuild.cmds bash -c "cmd"
+echo "HMM building completed."
+echo ""
+echo "Starting HMM search against other proteome..."
+mkdir -p search
+echo -n >hmmsearch.cmds
+for i in hmms/*.hmm; do
+    j=$(basename "$i")
+    echo "hmmsearch --tblout search/${j}search.tblout $i $proteome > search/${j}search.rawout" >>hmmsearch.cmds
+done
+xargs -t -P "$threads" -I cmd -a hmmsearch.cmds bash -c "cmd"
+echo "HMM search completed."
+echo ""
+echo "All steps completed successfully."
--- a/99.scripts/workflow/orthology_inference/01.orthofinder_sc_align.sh
+++ b/99.scripts/workflow/orthology_inference/01.orthofinder_sc_align.sh
@ -1,8 +0,0 @@
-#! /usr/bin/env bash
-mkdir -p msa
-echo -n > mafft.cmds
-for i in ogs/*.fa ; do
-    j=$(basename "$i")
-    echo "linsi --quiet $i > msa/$j" >> mafft.cmds
-done
-xargs -t -P 8 -I cmd -a mafft.cmds bash -c "cmd"
--- a/99.scripts/workflow/orthology_inference/02.hmmbuild.sh
+++ b/99.scripts/workflow/orthology_inference/02.hmmbuild.sh
@ -1,8 +0,0 @@
-#! /usr/bin/env bash
-mkdir -p hmms
-echo -n > hmmbuild.cmds
-for i in msa/*.fa ; do
-    j=$(basename "$i")
-    echo "hmmbuild -o hmms/${j}.hmmbuild.out --amino hmms/${j}.hmm $i" >> hmmbuild.cmds
-done
-xargs -t -P 8 -I cmd -a hmmbuild.cmds bash -c "cmd"
--- a/99.scripts/workflow/orthology_inference/02.macse_align.sh
+++ b/99.scripts/workflow/orthology_inference/02.macse_align.sh
@ -0,0 +1,34 @@
+#! /bin/bash
+set -e
+SCRIPTS=${SCRIPTS:-"$PROJECTHOME/99.scripts"}
+THREADS=${THREADS:-12}
+
+if [ "$#" -ne 5 ]; then
+    echo "Usage: $0 <ogs_dir> <hmmsearch_result_dir> <all_cds.fa> <output_dir> <homolog_stem>"
+    echo "Integrate hmmsearch results to new orthologous groups directory and perform MACSE alignment"
+    exit 1
+fi
+
+ogs_dir=$(readlink -f "$1")
+search_dir=$(readlink -f "$2")
+all_cds=$(readlink -f "$3")
+out_dir=$4
+stem=$5
+
+echo "Integrating hmmsearch results to new orthologous groups directory..."
+python3 "$SCRIPTS"/miscs/hmmsearch_result_to_new_ogs_dir.py \
+    -d "$ogs_dir" \
+    -t "$search_dir" \
+    -f "$all_cds" \
+    -o "$out_dir" \
+    -s "$stem"
+echo "Integration completed."
+
+echo "Starting MACSE alignment of orthologous groups..."
+echo -n >macse.cmds
+for og_dir in "$out_dir"/ogs/*; do
+    j=$(basename "$og_dir")
+    echo "cd $og_dir && bash $SCRIPTS/miscs/macse.sh ${j}_${stem}.fa ${j}.fa $j" >>macse.cmds
+done
+xargs -t -P "$THREADS" -I cmd -a macse.cmds bash -c "cmd"
+echo "MACSE alignment completed."
--- a/99.scripts/workflow/orthology_inference/03.hmmsearch.sh
+++ b/99.scripts/workflow/orthology_inference/03.hmmsearch.sh
@ -1,8 +0,0 @@
-#! /usr/bin/env bash
-mkdir -p hmmsearch
-echo -n > hmmsearch.cmds
-for i in hmms/*.hmm ; do
-    j=$(basename "$i")
-    echo "hmmsearch --tblout hmmsearch/${j}search.tblout $i ../../01.reference/Zju.pep.fa > hmmsearch/${j}search.rawout" >> hmmsearch.cmds
-done
-xargs -t -P 8 -I cmd -a hmmsearch.cmds bash -c "cmd"
--- a/99.scripts/workflow/orthology_inference/04.pep_align.sh
+++ b/99.scripts/workflow/orthology_inference/04.pep_align.sh
@ -1,8 +0,0 @@
-#! /usr/bin/env bash
-mkdir -p pep_aln
-echo -n > mafft.cmds
-for i in raw_ogs/pep/*.fa; do
-    j=$(basename "$i")
-    echo "linsi --quiet $i > pep_aln/${j/.fa/.pal}" >> mafft.cmds
-done
-xargs -t -P 8 -I cmd -a mafft.cmds bash -c "cmd"
--- a/99.scripts/workflow/orthology_inference/05.pal2nal.sh
+++ b/99.scripts/workflow/orthology_inference/05.pal2nal.sh
@ -1,8 +0,0 @@
-#! /usr/bin/env bash
-mkdir -p cds_aln
-echo -n > pal2nal.cmds
-for i in pep_aln/*.pal; do
-    j=$(basename "$i")
-    echo "pal2nal.pl $i raw_ogs/cds/${j/.pal/.fa} -output fasta > cds_aln/${j/.pal/.nal}" >> pal2nal.cmds
-done
-xargs -t -P 8 -I cmd -a pal2nal.cmds bash -c "cmd"
--- a/99.scripts/workflow/phylogeny_reconstruction/04.concatenate_ogs.py
+++ b/99.scripts/workflow/phylogeny_reconstruction/04.concatenate_ogs.py
@ -94,11 +94,18 @@ def concatenate_fasta_files(fasta_files, output_file):
    print(f"Total output sequence length: {len(concatenated_sequences[0].seq)}.")


-def get_fasta_files_from_directory(directory, extensions):
+def get_fasta_files_from_directory(directory, extensions, list_file=None):
    """
    get all FASTA files from a directory with specified extensions
    """
    fasta_files = []
+    if list_file:
+        with open(list_file, "r") as lf:
+            for line in lf:
+                filepath = os.path.join(directory, line.strip())
+                if os.path.isfile(filepath):
+                    fasta_files.append(filepath)
+        return sorted(fasta_files)
    for filename in os.listdir(directory):
        if any(filename.endswith(ext) for ext in extensions):
            fasta_files.append(os.path.join(directory, filename))
@ -119,12 +126,15 @@ def main():
        default=[".fasta", ".fa", ".fna"],
        help="FASTA file extensions to look for in directory",
    )
+    parser.add_argument("-l", "--list", help="List of input FASTA files", default=None)

    args = parser.parse_args()

    # 获取输入文件
    if args.directory:
-        fasta_files = get_fasta_files_from_directory(args.directory, args.extensions)
+        fasta_files = get_fasta_files_from_directory(
+            args.directory, args.extensions, args.list
+        )
        if not fasta_files:
            print(
                f"Cannot find FASTA files in {args.directory} with extensions {args.extensions}"
--- a/99.scripts/workflow/plastid/mapping.sh
+++ b/99.scripts/workflow/plastid/mapping.sh
@ -0,0 +1,15 @@
+#! /usr/bash
+if [ "$#" -ne 5 ]; then
+    echo "Usage: $0 <reference_genome> <fastq_1> <fastq_2> <output_directory> <stem>"
+    exit 1
+fi
+ref=$1
+fq1=$2
+fq2=$3
+outdir=$4
+stem=$5
+mkdir -p "$outdir"
+hisat -p 4 --dta -x "$ref" -1 "$fq1" -2 "$fq2" -S "${outdir}/${stem}.sam"
+samtools view -b -@ 4 "${outdir}/${stem}.sam" | samtools sort -@ 4 -o "${outdir}/${stem}.sorted.bam" --write-index
+rm "${outdir}/${stem}.sam"
+echo "Mapping completed. Sorted BAM file is at ${outdir}/${stem}.sorted.bam"
--- a/99.scripts/workflow/transcripts_assembly/01.denove_assembly.sh
+++ b/99.scripts/workflow/transcripts_assembly/01.denove_assembly.sh
@ -0,0 +1,31 @@
+#! /bin/bash
+set -e
+SCRIPTS=${SCRIPTS:-"$PROJECTHOME/99.scripts"}
+MAX_MEMORY=${MAX_MEMORY:-"20G"}
+VIRIDI=${VIRIDI:-"$PROJECTHOME/01.reference/viridiplantae_odb12/"}
+ROSALES=${ROSALES:-"$PROJECTHOME/01.reference/rosales_odb12/"}
+
+if [ "$#" -ne 4 ]; then
+    echo "Usage: $0 <reads_1.fastq> <reads_2.fastq> <stem> <threads>"
+    echo "Perform de novo transcriptome assembly using Trinity"
+    exit 1
+fi
+
+fq1=$1
+fq2=$2
+stem=$3
+outdir="$stem"_trinity_out_dir
+THREADS=$4
+
+
+# Run Trinity for de novo transcriptome assembly
+Trinity --seqType fq --left "$fq1" --right "$fq2" --CPU "$THREADS" --max_memory "$MAX_MEMORY" --output "$outdir"
+# Get Longest isoform per gene
+perl "$SCRIPTS"/trinity_utils/util/misc/get_longest_isoform_seq_per_trinity_gene.pl "$outdir.Trinity.fasta" >"$outdir".longest_isoform.fasta
+# BUSCO assessment
+busco -i "$outdir".longest_isoform.fasta -l "$VIRIDI" -m tran --cpu "$THREADS" -o "$outdir"_busco_viridi -f
+busco -i "$outdir".longest_isoform.fasta -l "$ROSALES" -m tran --cpu "$THREADS" -o "$outdir"_busco_rosales -f
+# Length Statistics
+TrinityStats.pl "$outdir.Trinity.fasta" >"$outdir".Trinity.fasta.length_stat.txt
+# Clear temporary directory
+rm -rf "$outdir"
--- a/99.scripts/workflow/transcripts_assembly/02.referenced_assembly.sh
+++ b/99.scripts/workflow/transcripts_assembly/02.referenced_assembly.sh
@ -0,0 +1,37 @@
+#! /bin/bash
+set -e
+MAX_MEMORY=${MAX_MEMORY:-"50G"}
+VIRIDI=${VIRIDI:-"$PROJECTHOME/01.reference/viridiplantae_odb12/"}
+ROSALES=${ROSALES:-"$PROJECTHOME/01.reference/rosales_odb12/"}
+SCRIPTS=${SCRIPTS:-"$PROJECTHOME/99.scripts"}
+
+if [ "$#" -ne 5 ]; then
+    echo "Usage: $0 <reads_1.fastq> <reads_2.fastq> <ref> <stem> <threads>"
+    echo "Perform reference-guided transcriptome assembly using Hisat2 and Trinity"
+    exit 1
+fi
+
+fq1=$1
+fq2=$2
+ref=$3
+stem=$4
+outdir="$stem"_trinity_out_dir
+THREADS=$5
+
+
+# Run Hisat2 for reads mapping to reference genome
+hisat2 -p "$THREADS" --dta -x "$ref" -1 "$fq1" -2 "$fq2" -S "$stem".sam
+samtools view -b -@ "$THREADS" -o "$stem".raw.bam "$stem".sam
+samtools sort -@ "$THREADS" -o "$stem".sorted.bam "$stem".raw.bam
+samtools index "$stem".sorted.bam
+rm "$stem".sam "$stem".raw.bam
+# Run Trinity for de novo transcriptome assembly
+Trinity --genome_guided_bam "$stem".sorted.bam --genome_guided_max_intron 10000 --max_memory "$MAX_MEMORY" --CPU "$THREADS" --output "$outdir"
+# Get Longest isoform per gene
+perl "$SCRIPTS"/trinity_utils/util/misc/get_longest_isoform_seq_per_trinity_gene.pl "$outdir/Trinity-GG.fasta" >"$outdir/longest_isoform.fasta"
+# BUSCO assessment
+busco -i "$outdir"/longest_isoform.fasta -l "$VIRIDI" -m tran --cpu "$THREADS" -o "$outdir"/busco_viridi -f
+busco -i "$outdir"/longest_isoform.fasta -l "$ROSALES" -m tran --cpu "$THREADS" -o "$outdir"/busco_rosales -f
+# Length Statistics
+TrinityStats.pl "$outdir"/Trinity-GG.fasta >"$outdir"/length_stat.txt
+
--- a/99.scripts/workflow/transcripts_assembly/03.td2_cds_predict.sh
+++ b/99.scripts/workflow/transcripts_assembly/03.td2_cds_predict.sh
@ -0,0 +1,18 @@
+#! /bin/bash
+set -e
+TMP=${TMP:-"$PROJECTHOME/tmp"}
+
+if [ "$#" -ne 3 ]; then
+    echo "Usage: $0 <transcripts_fasta> <swissprot_database> <output_directory>"
+    echo "Predict coding sequences (CDS) from transcripts using TD2 and MMseqs2"
+    exit 1
+fi
+transcripts=$1
+sprot=$2
+outdir=$3
+
+mkdir -p "$outdir"
+TD2.LongOrfs -t "$transcripts" --precise -@ 8 -O "$outdir"
+mmseqs easy-search "$outdir/longest_orfs.pep" "$sprot" "$outdir/mmseqs.m8" "$TMP" -s 7.0 --threads 16
+TD2.Predict -t "$transcripts" --precise -O "$outdir" --retain-mmseqs-hits "$outdir/mmseqs.m8"
+echo "CDS prediction completed. Results are in $outdir"
--- a/99.scripts/workflow/transcripts_assembly/04.longest_cds_rename.sh
+++ b/99.scripts/workflow/transcripts_assembly/04.longest_cds_rename.sh
@ -0,0 +1,25 @@
+#! /bin/bash
+set -e
+SCRIPTS=${SCRIPTS:-"$PROJECTHOME/99.scripts"}
+
+if [ "$#" -ne 3 ]; then
+    echo "Usage: $0 <input_dir> <output_dir> <extension>"
+    echo "Extract longest isoform per gene and rename sequences"
+    exit 1
+fi
+
+indir=$1
+outdir=$2
+ext=$3
+
+mkdir -p "$outdir"
+# Process each file in the input directory with the specified extension
+for td_cds in "$indir"/*."$ext"; do
+    stem=$(basename "$td_cds" ."$ext")
+    echo "Processing $td_cds($stem) ..."
+    echo "perl $SCRIPTS/trinity_utils/util/misc/get_longest_isoform_seq_per_trinity_gene.pl $td_cds > $outdir/$stem.longest_isoform.fa"
+    perl "$SCRIPTS"/trinity_utils/util/misc/get_longest_isoform_seq_per_trinity_gene.pl "$td_cds" >"$outdir"/"$stem".longest_isoform.fa
+    echo "$SCRIPTS/rename_trinity_fasta.py $outdir/$stem.longest_isoform.fa $stem $outdir/$stem.full_cds.fa"
+    "$SCRIPTS"/miscs/rename_trinity_fasta.py "$outdir"/"$stem".longest_isoform.fa "$stem" "$outdir"/"$stem".full_cds.fa
+    echo "Done."
+done
--- a/99.scripts/workflow/transcripts_assembly/05.reduce_redundancy.sh
+++ b/99.scripts/workflow/transcripts_assembly/05.reduce_redundancy.sh
@ -0,0 +1,27 @@
+#! /bin/bash
+set -e
+SCRIPTS=${SCRIPTS:-"$PROJECTHOME/99.scripts"}
+IDENTITY=${IDENTITY:-0.99}
+THREADS=${THREADS:-6}S
+
+if [ "$#" -ne 3 ]; then
+    echo "Usage: $0 <input_dir> <output_dir> <extension>"
+    echo "Reduce redundancy of CDS files using cd-hit-est"
+    exit 1
+fi
+
+indir=$1
+outdir=$2
+ext=$3
+
+mkdir -p "$outdir"
+# Process each file in the input directory with the specified extension
+for cds in "$indir"/*."$ext"; do
+    stem=$(basename "$cds" ."$ext")
+    echo "Processing $cds($stem) ..."
+    echo "cd-hit-est -i $cds -o $outdir/$stem.cds_rr.fa -c $IDENTITY -n 10 -r 0 -T $THREADS"
+    cd-hit-est -i "$cds" -o "$outdir/$stem".cds_rr.fa -c "$IDENTITY" -n 10 -r 0 -T "$THREADS"
+    echo "seqkit translate $outdir/$stem.cds_rr.fa > $outdir/$stem.prot_rr.fa"
+    seqkit translate "$outdir/$stem".cds_rr.fa >"$outdir/$stem".prot_rr.fa
+    echo "Done."
+done
--- a/pixi.lock
+++ b/pixi.lock
@ -12,6 +12,7 @@ environments:
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/_openmp_mutex-4.5-2_gnu.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/_r-mutex-1.0.1-anacondar_1.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/alsa-lib-1.2.14-hb9d3cd8_0.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/aom-3.9.1-hac33072_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/archspec-0.2.5-pyhd8ed1ab_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/argcomplete-3.6.3-pyhd8ed1ab_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/aria2-1.37.0-hbc8128a_2.conda
@ -94,9 +95,11 @@ environments:
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/contourpy-1.3.3-py311hdf67eae_3.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/coreutils-9.5-hd590300_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/cpp-expected-1.3.1-h171cf75_0.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/cpython-3.11.14-py311hd8ed1ab_2.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/curl-8.17.0-h4e3cde8_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/cycler-0.12.1-pyhd8ed1ab_1.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/cyrus-sasl-2.1.28-hd9c7081_0.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/dav1d-1.2.1-hd590300_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/dbus-1.16.2-h3c4dab8_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/noarch/dendropy-5.0.8-pyhdfd78af_1.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/linux-64/diamond-2.1.16-h13889ed_0.conda
@ -109,6 +112,7 @@ environments:
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/linux-64/fastme-2.1.6.3-h7b50bb2_1.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/linux-64/fastp-1.0.1-heae3180_0.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/linux-64/fasttree-2.2.0-h7b50bb2_0.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/filelock-3.20.0-pyhd8ed1ab_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/fmt-11.2.0-h07f6e7f_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/font-ttf-dejavu-sans-mono-2.37-hab24e00_0.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/font-ttf-inconsolata-3.000-h77eed37_0.tar.bz2
@ -121,6 +125,7 @@ environments:
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/freetype-2.14.1-ha770c72_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/fribidi-1.0.16-hb03c661_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/frozendict-2.4.7-py311h49ec1c0_0.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/fsspec-2025.12.0-pyhd8ed1ab_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gawk-5.3.1-hcd3d067_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gcc_impl_linux-64-15.2.0-hcacfade_7.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gettext-0.25.1-h3f43e3d_1.conda
@ -132,6 +137,7 @@ environments:
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/glib-tools-2.86.1-hf516916_2.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/glpk-5.0-h445213a_0.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gmp-6.3.0-hac33072_2.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gmpy2-2.2.1-py311h92a432a_2.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/graphite2-1.3.14-hecca717_2.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gsl-2.7-he838d99_0.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/gst-plugins-base-1.24.11-h651a532_0.conda
@ -149,12 +155,14 @@ environments:
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/idna-3.11-pyhd8ed1ab_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/linux-64/iqtree-3.0.1-h503566f_0.tar.bz2
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/isa-l-2.31.1-hb9d3cd8_1.conda
+      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/jinja2-3.1.6-pyhcf101f3_1.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/joblib-1.5.2-pyhd8ed1ab_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/jq-1.8.1-h73b1eb8_0.conda
      - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/jsoncpp-1.9.6-hf42df4d_1.conda
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  - tbb 2022.3.0 h8d10470_1
  size: 1115083
  timestamp: 1762509972811
+- conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/bioconda/noarch/td2-1.0.6-pyhdfd78af_0.tar.bz2
+  sha256: e361bb3ba0ed7d1f9a4d8423058763c50e00c2e964d19a6491e09e903345987d
+  md5: b3a676270ab795aad86bdceb859f4128
+  depends:
+  - numpy >=1.24.4,<2
+  - pandas >=2.0.3
+  - psauron >=1.0.5
+  - psutil
+  - python >=3.9,<3.13
+  - pytorch >=2.1.2
+  - scipy >=1.10.1
+  - setuptools <81
+  - torchaudio >=2.1.2
+  - torchvision >=0.16.2
+  - tqdm >=4.66.1
+  - typing_extensions >=4.9.0
+  license: MIT
+  license_family: MIT
+  size: 45099
+  timestamp: 1749493915044
 - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/threadpoolctl-3.6.0-pyhecae5ae_0.conda
  sha256: 6016672e0e72c4cf23c0cf7b1986283bd86a9c17e8d319212d78d8e9ae42fdfd
  md5: 9d64911b31d57ca443e9f1e36b04385f
@ -9036,6 +9526,72 @@ packages:
  license_family: MIT
  size: 38777
  timestamp: 1749127286558
+- conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/torchaudio-2.7.0-cpu_py311h1666e02_0.conda
+  sha256: eabf0989faf51ba30eabeb9256d9f4ef48d0af5a84b88d0baafe2c8fdb941ef6
+  md5: 32ac754bca22e7a23d18cb3792b2112b
+  depends:
+  - python
+  - numpy
+  - kaldi
+  - pytorch * cpu*
+  - __glibc >=2.17,<3.0.a0
+  - libgcc >=13
+  - libstdcxx >=13
+  - libgcc >=13
+  - libzlib >=1.3.1,<2.0a0
+  - pytorch >=2.6.0,<2.7.0a0
+  - libtorch >=2.6.0,<2.7.0a0
+  - kaldi >=5.5.1112,<5.5.1113.0a0
+  - python_abi 3.11.* *_cp311
+  - liblzma >=5.8.1,<6.0a0
+  - bzip2 >=1.0.8,<2.0a0
+  constrains:
+  - llvmlite >=0.44
+  license: BSD-2-Clause
+  license_family: BSD
+  size: 4689009
+  timestamp: 1745964368346
+- conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/torchvision-0.21.0-cpu_py311_hfa9c634_1.conda
+  sha256: a5971392370b8f7755118e72bd40ec3bdc026ea59b8237d848842a4414170ab7
+  md5: af5a1b1a68a53f27bc3c87ab7c582e6d
+  depends:
+  - python
+  - pytorch * cpu*
+  - pillow >=5.3.0,!=8.3.0,!=8.3.1
+  - numpy >=1.23.5
+  - torchvision-extra-decoders
+  - __glibc >=2.17,<3.0.a0
+  - libgcc >=13
+  - libstdcxx >=13
+  - libgcc >=13
+  - libpng >=1.6.47,<1.7.0a0
+  - libwebp-base >=1.5.0,<2.0a0
+  - pytorch >=2.6.0,<2.7.0a0
+  - libtorch >=2.6.0,<2.7.0a0
+  - libjpeg-turbo >=3.0.0,<4.0a0
+  - giflib >=5.2.2,<5.3.0a0
+  - python_abi 3.11.* *_cp311
+  license: BSD-3-Clause
+  license_family: BSD
+  size: 1457950
+  timestamp: 1742619225483
+- conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/torchvision-extra-decoders-0.0.2-py311h896098c_2.conda
+  sha256: 424417d1fa2c0b1c5eaf218b71b398522cc9082f2df2bb8f88921731c1bc45e7
+  md5: aaebf7b5393e30ed452de175834ff032
+  depends:
+  - python
+  - libstdcxx >=13
+  - libgcc >=13
+  - __glibc >=2.17,<3.0.a0
+  - libheif >=1.19.5,<1.20.0a0
+  - python_abi 3.11.* *_cp311
+  - pytorch >=2.6.0,<2.7.0a0
+  - libtorch >=2.6.0,<2.7.0a0
+  - libavif16 >=1.1.1,<2.0a0
+  - libtorch >=2.6.0,<2.7.0a0
+  license: LGPL-2.1-only
+  size: 65750
+  timestamp: 1739805887089
 - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/tornado-6.5.2-py311h49ec1c0_2.conda
  sha256: 1913516458f92df2a0b415426dce27cc14922415787f4b672a707b233631b1e0
  md5: 8d7a63fc9653ed0bdc253a51d9a5c371
@ -9152,6 +9708,15 @@ packages:
  license_family: MIT
  size: 23801
  timestamp: 1753886790616
+- conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/typing-extensions-4.15.0-h396c80c_0.conda
+  sha256: 7c2df5721c742c2a47b2c8f960e718c930031663ac1174da67c1ed5999f7938c
+  md5: edd329d7d3a4ab45dcf905899a7a6115
+  depends:
+  - typing_extensions ==4.15.0 pyhcf101f3_0
+  license: PSF-2.0
+  license_family: PSF
+  size: 91383
+  timestamp: 1756220668932
 - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/noarch/typing_extensions-4.15.0-pyhcf101f3_0.conda
  sha256: 032271135bca55aeb156cee361c81350c6f3fb203f57d024d7e5a1fc9ef18731
  md5: 0caa1af407ecff61170c9437a808404d
@ -9259,6 +9824,16 @@ packages:
  license_family: MIT
  size: 62931
  timestamp: 1733130309598
+- conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/x265-3.5-h924138e_3.tar.bz2
+  sha256: 76c7405bcf2af639971150f342550484efac18219c0203c5ee2e38b8956fe2a0
+  md5: e7f6ed84d4623d52ee581325c1587a6b
+  depends:
+  - libgcc-ng >=10.3.0
+  - libstdcxx-ng >=10.3.0
+  license: GPL-2.0-or-later
+  license_family: GPL
+  size: 3357188
+  timestamp: 1646609687141
 - conda: https://mirrors.tuna.tsinghua.edu.cn/anaconda/cloud/conda-forge/linux-64/xcb-util-0.4.1-h4f16b4b_2.conda
  sha256: ad8cab7e07e2af268449c2ce855cbb51f43f4664936eff679b1f3862e6e4b01d
  md5: fdc27cb255a7a2cc73b7919a968b48f0
--- a/pixi.toml
+++ b/pixi.toml
@ -52,6 +52,8 @@ ete3 = ">=3.1.3,<4"
 xvfbwrapper = ">=0.2.15,<0.3"
 r-ggplot2 = ">=4.0.1,<5"
 macse = ">=2.7,<3"
+td2 = ">=1.0.6,<2"
+mmseqs2 = ">=18.8cc5c,<19"

 [feature.mrbayes.dependencies]
 beagle-lib = ">=3.1.2,<4"
--- a/slides/.gitignore
+++ b/slides/.gitignore
@ -0,0 +1,7 @@
+node_modules
+.DS_Store
+dist
+*.local
+.vite-inspect
+.remote-assets
+components.d.ts
--- a/slides/.npmrc
+++ b/slides/.npmrc
@ -0,0 +1,3 @@
+# for pnpm
+shamefully-hoist=true
+auto-install-peers=true
--- a/slides/20251213/pages/审稿意见与解决方案.md
+++ b/slides/20251213/pages/审稿意见与解决方案.md
@ -0,0 +1,136 @@
+---
+layout: section
+color: emerald
+---
+
+<div class="w-7/10 mx-auto text-center">
+
+## 审稿意见与解决方案
+
+<hr/>
+
+仅列出涉及到流程或数据方面的审稿意见及解决方案
+
+</div>
+
+---
+layout: top-title
+color: emerald
+title: 方法学描述不清与创新性未突出
+
+---
+
+::title::
+
+# 方法学描述不清与创新性未突出
+
+::content::
+
+- **核心问题**：所提出的新方法（基于MCMC拟合单峰/双峰分布推断次级分歧时间）缺乏清晰、系统的阐述。
+- **具体建议**：
+  - 明确说明新方法相较于现有方法（如 D3 test, PhyloNet 等）的**主要优势**（例如是否能同时识别introgressed基因并估计introgression发生时间？）。
+  - 在**摘要、引言、方法、讨论**各部分突出方法的创新点，而非仅模糊称为“MCMC拟合模型”。
+  - 澄清关键术语与参数含义（如 Line 320/356 中的 “r” 是什么速率？Line 353 的“偏差”指什么？）。
+  - 解释“组合比例”（combination ratio）和“分类因子”（classification factor）如何计算或估计。
+  - 说明用于判断单峰 vs. 双峰分布优劣的**统计检验方法**（如 AIC、BIC、Bayes因子等）。
+
+- **解决方案：**
+
+  - 论文文字修改：模型描述改进
+  - ==开发分布优劣的统计检验方法==
+
+---
+layout: top-title
+color: emerald
+title: 方法验证不足
+
+---
+
+::title::
+
+# 方法验证不足
+
+::content::
+
+- **核心问题**：仅在一个小型植物类群（Hippophae，10个种）中应用，缺乏普适性验证。
+- **具体建议**：
+  - 增加**模拟数据实验**，验证方法在不同introgression强度、重复事件、ILS水平下的表现。
+  - 或至少在**其他真实数据集**（不同类群）中测试方法，证明其可推广性。
+  - 讨论方法的**适用范围与局限性**（如是否适用于大规模物种？能否处理多次introgression？）。
+- **解决方案：**
+  - ==bpp模拟数据实验==
+  - ==添加其他真实数据集模拟（蛇）==
+
+---
+layout: top-title
+color: emerald
+title: 系统发育与分化时间推断存在潜在偏差
+
+---
+
+::title::
+
+# 系统发育与分化时间推断存在潜在偏差
+
+::content::
+
+- **核心问题**：分化时间估计可能受取样策略和模型选择影响。
+- **具体建议**：
+
+  - 检查是否所有基因树具有**一致的分类单元取样**；若不一致，需说明如何校正由此带来的年龄估计偏差。
+  - 质疑使用**Yule过程**作为先验（尤其当包含种内样本与远缘类群时），建议考虑更合适的物种形成模型。
+  - 说明**核苷酸替代模型（如HKY）的选择依据**，是否进行过模型选择（如jModelTest, ModelFinder等）。
+  - 澄清图1与图4中**分化时间估计差异的原因**，评估其对introgression时间推断的影响。
+- **解决方案：**
+
+  - ==使用MCMCTree替代beast做分化时间计算，可以直接使用更好的替代模型==
+  - ==使用少量基因验证yule过程作为物种形成模型的合理性==
+  - 论文文字修改：增加对基因树取样一致性和物种形成模型选择的讨论
+
+---
+layout: top-title
+color: emerald
+title: 数据处理与结果解释需加强严谨性
+
+---
+
+::title::
+
+# 数据处理与结果解释需加强严谨性
+
+::content::
+
+- **核心问题**：从3149个单拷贝基因中仅保留177个“可靠”基因，可能导致结果偏倚。
+- **具体建议**：
+
+  - 明确“可靠基因”的筛选标准（高bootstrap支持 ≠ 正确），建议改称“informative genes”。
+  - 强调所识别的64个introgressed基因**不代表全基因组水平的introgression比例**，因多拷贝或功能重要基因可能被过滤掉。
+  - 对这64个基因进行**功能注释分析**，探讨其生物学意义。
+  - 排除**污染**作为introgression信号来源的可能性（如通过检查reads mapping、组织来源等）。
+- **解决方案：**
+
+  - ==探索模型是否可能涵盖低支持度的基因，并在后续评估informative genes结果是否与之相近，能否替代==
+
+---
+layout: top-title
+color: emerald
+title: 材料与可重复性问题
+
+---
+
+::title::
+
+# 材料与可重复性问题
+
+::content::
+
+- **核心问题**：缺乏关键实验细节和代码共享。
+- **具体建议**：
+
+  - 提供**新方法的完整代码**（如GitHub链接），确保他人可复现。
+  - 补充**凭证标本信息**（采集号、保存的标本馆）。
+  - 将原始测序数据（raw reads）提交至**SRA**，而非作为补充材料；补充材料应包含组装后的转录本、CDS、蛋白序列等，并拆分为小于1GB的压缩包。
+- **解决方案：**
+
+  - ==从头分析，备份完整代码与中间文件==
+  - ==使用rstan重新实现MCMC模型，方便提交代码==
--- a/slides/20251213/pages/技术路线.md
+++ b/slides/20251213/pages/技术路线.md
@ -0,0 +1,150 @@
+---
+layout: section
+color: navy
+---
+
+<div class="w-7/10 mx-auto text-center">
+
+## 技术路线与分析进展
+
+<hr/>
+
+当前分析进展情况
+
+</div>
+
+---
+layout: side-title
+color: navy
+align: rm-lm
+titlewidth: is-3
+title: 技术路线图（上）
+
+---
+
+::title::
+
+# 技术路线图（上）
+
+::content::
+
+```mermaid
+flowchart TD
+
+classDef default fill:#ECF5FF,stroke:#409eff,stroke-width:2px,color:#409eff;
+classDef done fill:#F0F9EB,stroke:#67c23a,stroke-width:2px,color:#67c23a;
+classDef warn fill:#FDF6EC,stroke:#e6a23c,stroke-width:2px,color:#e6a23c
+
+subgraph subgraph1[Step1. 数据预处理与组装]
+    direction LR
+    RawData[原始数据，来自sra]:::warn --> CleanData[数据清洗与质控，Fastp]:::done
+    CleanData -- 沙棘样品，有参 --> AlignData[比对到参考基因组，Hisat2]:::done
+    AlignData --> Assembly[组装成转录本，Trinity]:::done
+    CleanData -- 胡颓子，无参 --> Assembly[组装成转录本，使用N50和busco评估组装结果，Trinity]:::done
+end
+
+subgraph subgraph2[Step2. 同源基因集聚类]
+    direction LR
+    ass[转录本]:::warn --> CDS[预测CDS，抽取最长转录本的最长ORF作为输入CDS，TransDecoder]:::done
+    CDS --> reducedCDS[单样本CDS序列去冗余，CD-HIT]:::done
+    reducedCDS --> ortho[基因家族聚类得到低拷贝同源基因集，OrthoFinder]:::done
+    ortho --> homolog[将枣的同源基因整合到低拷贝同源基因集中, hmmer]:::done
+end
+
+subgraph subgraph3[Step3. 基因集筛选与基因树]
+    direction LR
+    homo[低拷贝同源基因集]:::warn --> alignment[基于蛋白序列比对，mafft/pal2nal<br>修剪比对序列，trimAl]:::done
+    alignment --> treeshrink[fasttree快速构树，基于长枝吸引（treeshrink）和比对序列长度过滤基因集]:::done
+    treeshrink --> modeltest[序列进化模型检验，modeltest-ng]:::done
+    modeltest --> raxml[构建ML单基因树]:::done
+    modeltest --> mrbayes[构建BP单基因树]:::done
+    raxml --> trees[根据mrbayes的结果收敛性筛选基因树与对应序列]:::done
+    mrbayes --> trees
+end
+
+subgraph subgraph4[Step4. 物种系统发育关系]
+    direction LR
+    tree[过滤后单基因树与对应序列]:::warn -- 单基因ML树 --> coalescence[溯祖关系树, Aster]
+    tree -- 单基因ML树 --> phyparts[基因树不一致性展示]
+    tree -- 单基因序列 --> concatenation[串联关系树，iqtree3]
+    tree -- 单基因的mrbayes分析结果 --> CFs[基因树一致性因子，BUCKy]:::done
+    CFs --> network[系统发育网络，SNAQ]:::done
+    coalescence --> network
+end
+
+subgraph note[节点说明]
+    direction TB
+    done[已完成分析]:::done
+    warn[重要中间数据]:::warn
+    default[未完成分析]
+end
+
+subgraph1 --> subgraph2
+subgraph2 --> subgraph3
+subgraph3 --> subgraph4
+subgraph4 ~~~ note
+```
+
+---
+layout: side-title
+color: navy
+align: rm-lm
+titlewidth: is-3
+title: 技术路线图（下）
+
+---
+
+::title::
+# 技术路线图（下）
+
+::content::
+
+```mermaid
+flowchart TD
+
+classDef default fill:#ECF5FF,stroke:#409eff,stroke-width:2px,color:#409eff;
+classDef done fill:#F0F9EB,stroke:#67c23a,stroke-width:2px,color:#67c23a;
+classDef warn fill:#FDF6EC,stroke:#e6a23c,stroke-width:2px,color:#e6a23c
+
+subgraph subgraph5[Step5. 叶绿体系统发育关系]
+    direction LR
+    cleanData[Clean Reads]:::done --> alignCp[比对到参考叶绿体基因组，hisat2]:::done
+    alignCp --> callExon[提取比对上的外显子序列，samtools]
+    callExon --> cpTree[串联高质量的exon片段，构建叶绿体基因组系统发育树，iqtree3]
+    cpTree --> divergence[估算叶绿体基因组分歧时间，beast]
+end
+
+subgraph subgraph6[Step6. 沙棘模型分析]
+    direction LR
+    genes[筛选得到的低拷贝同源基因集]:::warn --> filterGenes[基于ml树筛选基因集用于后续模型]
+    filterGenes --> mcmcTree[使用mcmctree估算分化时间]
+    mcmcTree --> fitModel[使用模型拟合种间分化时间分布]
+    fitModel --> integrate[整合不同模型结果得到最终分化时间估计]
+end
+
+subgraph subgraph7[Step7. bpp模拟模型分析]
+    direction LR
+    bpp[使用bpp进行溯祖模拟得到基因集]:::warn --> mcmcTreeBPP[使用mcmctree估算分化时间]
+    mcmcTreeBPP --> fitModelBPP[使用模型拟合种间分化时间分布]
+    fitModelBPP --> integrateBPP[整合不同模型结果得到最终分化时间估计]
+end
+
+subgraph subgraph8[Step8. 蛇数据分析]
+    direction LR
+    snakeData[蛇数据]:::warn --> mcmcTreeSnake[使用mcmctree估算分化时间]
+    mcmcTreeSnake --> fitModelSnake[使用模型拟合种间分化时间分布]
+    fitModelSnake --> integrateSnake[整合不同模型结果得到最终分化时间估计]
+end
+
+subgraph note[节点说明]
+    direction TB
+    done[已完成分析]:::done
+    warn[重要中间数据]:::warn
+    default[未完成分析]
+end
+
+subgraph5 --> subgraph6
+subgraph6 --> subgraph7
+subgraph7 --> subgraph8
+subgraph8 ~~~ note
+```
--- a/slides/20251213/pages/结果.md
+++ b/slides/20251213/pages/结果.md
--- a/slides/20251213/slides.md
+++ b/slides/20251213/slides.md
@ -0,0 +1,23 @@
+---
+theme: neversink
+title: 论文修改进展|2025.12.13
+colorSchema: light
+layout: cover
+routerMode: hash
+transition: fade
+---
+
+# 论文修改进展
+
+#### 2025.12.13
+
+---
+src: ./pages/审稿意见与解决方案.md
+title: 审稿意见与解决方案
+---
+
+---
+src: ./pages/技术路线.md
+title: 技术路线与分析进展
+
+---
--- a/slides/netlify.toml
+++ b/slides/netlify.toml
@ -0,0 +1,16 @@
+[build]
+publish = "dist"
+command = "npm run build"
+
+[build.environment]
+NODE_VERSION = "20"
+
+[[redirects]]
+from = "/.well-known/*"
+to = "/.well-known/:splat"
+status = 200
+
+[[redirects]]
+from = "/*"
+to = "/index.html"
+status = 200
--- a/slides/package.json
+++ b/slides/package.json
@ -0,0 +1,17 @@
+{
+  "name": "slides",
+  "type": "module",
+  "private": true,
+  "scripts": {
+    "build": "slidev build",
+    "dev": "slidev --open",
+    "export": "slidev export"
+  },
+  "dependencies": {
+    "@slidev/cli": "^52.11.0",
+    "@slidev/theme-default": "latest",
+    "@slidev/theme-seriph": "latest",
+    "slidev-theme-neversink": "^0.4.0",
+    "vue": "^3.5.25"
+  }
+}
--- a/slides/pnpm-lock.yaml
+++ b/slides/pnpm-lock.yaml
--- a/slides/vercel.json
+++ b/slides/vercel.json
@ -0,0 +1,7 @@
+{
+  "rewrites": [
+    { "source": "/(.*)", "destination": "/index.html" }
+  ],
+  "buildCommand": "npm run build",
+  "outputDirectory": "dist"
+}