offtracker 2.12.2__zip → 2.13.0__zip

{offtracker-2.12.2/offtracker.egg-info → offtracker-2.13.0}/PKG-INFO

@@ -1,6 +1,6 @@
 Metadata-Version: 2.1
 Name: offtracker
-Version: 2.12.2
+Version: 2.13.0
 Summary: Tracking-seq data analysis
 Home-page: https://github.com/Lan-lab/offtracker
 Author: Runda Xu

{offtracker-2.12.2 → offtracker-2.13.0}/offtracker/X_offtracker.py

@@ -2,7 +2,8 @@
 import pandas as pd
 import polars as pl
 import numpy as np
-import os, sys
+import os, sys, re
+import offtracker.X_sequence as xseq
 sys.path.append( os.path.abspath(os.path.dirname(__file__)) )
 
 def fdr(p_vals):
@@ -115,6 +116,29 @@ def target_signal(df_bdg_chr, chrom, cleavage_site, flank_max=100000, smooth_tim
                   binsize=100, flank_regions=[500,1000,2000,5000], 
                   length_bkg = 20000, length_binsize=1000, length_min_noise=0.2, n_std=1, 
                   end='end',start='start',value='residual', pct_offset=0.0):
+    """_summary_
+
+    Args:
+        df_bdg_chr (_type_): .bdg table with the same chromosome
+        chrom (_type_): chr name
+        cleavage_site (_type_): cleavage site
+        flank_max (int, optional): _description_. Defaults to 100000.
+        smooth_times (int, optional): _description_. Defaults to 1.
+        window_size (int, optional): _description_. Defaults to 3.
+        binsize (int, optional): _description_. Defaults to 100.
+        flank_regions (list, optional): _description_. Defaults to [500,1000,2000,5000].
+        length_bkg (int, optional): _description_. Defaults to 20000.
+        length_binsize (int, optional): _description_. Defaults to 1000.
+        length_min_noise (float, optional): _description_. Defaults to 0.2.
+        n_std (int, optional): _description_. Defaults to 1.
+        end (str, optional): _description_. Defaults to 'end'.
+        start (str, optional): _description_. Defaults to 'start'.
+        value (str, optional): _description_. Defaults to 'residual'.
+        pct_offset (float, optional): _description_. Defaults to 0.0.
+
+    Returns:
+        _type_: _description_
+    """
     # 输入数据必须是同一条染色体内的
     # 统计 flank regions 的个数
     # n_regions = len(flank_regions)
@@ -328,6 +352,25 @@ def target_signal(df_bdg_chr, chrom, cleavage_site, flank_max=100000, smooth_tim
 
 def target_signal_chunk(df_bdg_chr, df_alignment_chr, flank_max=100000, smooth_times = 1, window_size = 3, binsize=100, flank_regions=[500,1000,2000,5000], 
                         length_bkg = 20000, length_binsize=1000, length_min_noise=0.2, n_std=1, pct_offset=0.0):
+    """
+
+    Args:
+        df_bdg_chr (_type_): .bdg table with the same chromosome
+        df_alignment_chr (_type_): candidate sites
+        flank_max (int, optional): _description_. Defaults to 100000.
+        smooth_times (int, optional): _description_. Defaults to 1.
+        window_size (int, optional): _description_. Defaults to 3.
+        binsize (int, optional): _description_. Defaults to 100.
+        flank_regions (list, optional): _description_. Defaults to [500,1000,2000,5000].
+        length_bkg (int, optional): _description_. Defaults to 20000.
+        length_binsize (int, optional): _description_. Defaults to 1000.
+        length_min_noise (float, optional): _description_. Defaults to 0.2.
+        n_std (int, optional): _description_. Defaults to 1.
+        pct_offset (float, optional): _description_. Defaults to 0.0.
+
+    Returns:
+        _type_: _description_
+    """
     # 输入数据必须是同一条染色体内的
     list_target_all = []
     for a_row in df_alignment_chr.iterrows():
@@ -348,3 +391,261 @@ def target_signal_chunk(df_bdg_chr, df_alignment_chr, flank_max=100000, smooth_t
     return df_result
 
 
+
+
+
+
+
+
+
+
+
+############################################################################
+# 2025.08.08 新写的的基于 Bio.Align 的 local realign，用于局部修正脱靶位点坐标 #
+############################################################################
+def create_substitution_matrix(mismatch_score=0.01):
+    """
+    Create substitution matrix for DNA alignment using Bio.Align format.
+    """
+    alphabet = 'ATGCN'
+    matrix = np.full((len(alphabet), len(alphabet)), mismatch_score)
+    
+    # Set match scores
+    for i in range(len(alphabet)):
+        matrix[i][i] = 2.0
+    
+    # N matches with everything
+    n_idx = alphabet.index('N')
+    matrix[n_idx, :] = 2.0
+    matrix[:, n_idx] = 2.0
+    
+    return matrix, alphabet
+
+def sgRNA_alignment_new(a_key, sgRNA, seq, substitution_matrix=None, alphabet=None, 
+                   mismatch_score=0.01):
+    """
+    Perform local alignment using Bio.Align instead of deprecated pairwise2.
+    """
+    from Bio import Align
+    if substitution_matrix is None or alphabet is None:
+        substitution_matrix, alphabet = create_substitution_matrix(mismatch_score)
+    
+    # Create aligner
+    aligner = Align.PairwiseAligner()
+    aligner.substitution_matrix = Align.substitution_matrices.Array(
+        alphabet=alphabet, dims=2, data=substitution_matrix
+    )
+    aligner.open_gap_score = -2
+    aligner.extend_gap_score = -2
+    aligner.mode = 'local'
+    
+    try:
+        # Perform alignment
+        alignments = aligner.align(sgRNA, seq)
+
+        if not alignments:
+            # No alignment found, return default values
+            return [0, 0, '', f"{a_key.split(':')[0]}:0-0", 0, 0, len(sgRNA)]
+        
+        # Convert to list for indexing
+        alignments = list(alignments)
+         
+        # Extract alignment information
+        coords = alignments[0].coordinates
+        start_target = coords[1][0]
+        end_target = coords[1][-1]
+        
+        # Extract target sequence directly from coordinates
+        # target = seq[start_target:end_target]
+        
+        # Get aligned sequences for detailed analysis
+        alignment_str = str(alignments[0])
+        alignment_lines = alignment_str.split('\n')
+        if len(alignment_lines) >= 3:
+            aligned_sgrna = [x for x in alignment_lines[0].split(' ') if x != '']
+            aligned_genome = [x for x in alignment_lines[2].split(' ') if x != '']
+        else:
+            raise ValueError("Unexpected alignment format")
+        
+        assert int(aligned_sgrna[-1]) == len(sgRNA)
+        
+        # Calculate indels and mismatches
+        # deletion = RNA bulge
+        # insertion = DNA bulge
+        aligned_sgrna_seq = aligned_sgrna[-2]
+        aligned_genome_seq = aligned_genome[-2]
+        insertion = aligned_sgrna_seq.count('-') if '-' in aligned_sgrna_seq else 0
+        deletion = aligned_genome_seq.count('-') if '-' in aligned_genome_seq else 0
+        
+        # Count mismatches by comparing sequences directly
+        # mismatch = 0
+        # assert len(aligned_sgrna_seq) == len(aligned_genome_seq)
+        # for i in range(len(aligned_sgrna_seq)):
+        #     if (aligned_sgrna_seq[i] != aligned_genome_seq[i]) & (aligned_sgrna_seq[i] != 'N') & (aligned_genome_seq[i] != 'N'):
+        #         mismatch += 1
+
+        mismatch = round((alignments[0].score % 1)/mismatch_score)
+        
+        # Calculate target location
+        pos_st = int(a_key.split('-')[0].split(':')[1]) + 1
+        chr_name = a_key.split(':')[0]
+        target_st = pos_st + start_target
+        target_ed = pos_st + end_target - 1
+        target_location = f"{chr_name}:{target_st}-{target_ed}"
+        
+        score = alignments[0].score
+        
+        return [score, aligned_genome_seq, target_location, deletion, insertion, mismatch]
+            
+    except Exception as e:
+        print(f"Alignment error for {a_key}: {e}")
+        return [0, 0, '', f"{a_key.split(':')[0]}:0-0", 0, 0, len(sgRNA)]
+
+def local_realign(sgRNA_seq, fasta, PAM='NGG', PAM_loc='downstream'):
+    # 添加 PAM
+    if PAM_loc == 'downstream':
+        sgRNA_PAM_fw = sgRNA_seq + PAM
+    else:
+        sgRNA_PAM_fw = PAM + sgRNA_seq
+    sgRNA_PAM_rv = xseq.reverse_complement(sgRNA_PAM_fw)
+    list_args_fw=[]
+    list_args_rv=[]
+    for a_key, a_seq in fasta.items():
+        # 2025.04.25 修正大小写问题
+        a_seq = re.sub('[^ATCG]','N',a_seq.upper())
+        list_args_fw.append( [a_key, sgRNA_PAM_fw, a_seq])
+        list_args_rv.append( [a_key, sgRNA_PAM_rv, a_seq])
+    list_align_forward = [sgRNA_alignment_new(*args) for args in list_args_fw]
+    list_align_reverse = [sgRNA_alignment_new(*args) for args in list_args_rv]
+    #
+    df_align_forward = pd.DataFrame(list_align_forward, columns= ['fw_score', 'fw_target','fw_location','fw_deletion','fw_insertion','fw_mismatch'])
+    df_align_reverse = pd.DataFrame(list_align_reverse, columns= ['rv_score', 'rv_target','rv_location','rv_deletion','rv_insertion','rv_mismatch'])
+    df_align_reverse['rv_target'] = df_align_reverse['rv_target'].apply(xseq.reverse_complement)
+    df_candidate = pd.concat([df_align_forward,df_align_reverse],axis=1)
+    df_candidate['location'] = fasta.keys()
+    df_candidate['alignment_score'] = df_candidate[['fw_score','rv_score']].max(axis=1)
+    df_candidate['best_seq_score'] = df_candidate[['fw_score', 'rv_score']].max(axis=1)
+    df_candidate['best_strand'] = df_candidate[['fw_score', 'rv_score']].idxmax(axis='columns').replace({'fw_score':'+', 'rv_score':'-'})
+    df_candidate.loc[df_candidate['fw_score']==df_candidate['rv_score'],'best_strand']='equal_score'
+            
+    # GG check
+    # 2023.12.05 增加 cleavage_site 推测
+    list_best_target = []
+    list_best_location = []
+    list_cleavage_site = []
+    list_delete = []
+    list_insert = []
+    list_mismat = []
+    list_GG = []
+    for a_row in df_candidate.iterrows():
+        if a_row[1]['best_strand']=='+':
+            list_best_target.append(a_row[1]['fw_target'])
+            list_best_location.append(a_row[1]['fw_location'])
+            list_cleavage_site.append(int(a_row[1]['fw_location'].split('-')[1]) - 6)
+            list_delete.append(a_row[1]['fw_deletion'])
+            list_insert.append(a_row[1]['fw_insertion'])
+            list_mismat.append(a_row[1]['fw_mismatch'])
+            if a_row[1]['fw_target'][-2:]=='GG':
+                list_GG.append('OK')
+            else:
+                list_GG.append('NO')                     
+        elif a_row[1]['best_strand']=='-':
+            list_best_target.append(a_row[1]['rv_target'])
+            list_best_location.append(a_row[1]['rv_location'])
+            list_cleavage_site.append(int(a_row[1]['rv_location'].split('-')[0].split(':')[1]) + 5)
+            list_delete.append(a_row[1]['rv_deletion'])
+            list_insert.append(a_row[1]['rv_insertion'])
+            list_mismat.append(a_row[1]['rv_mismatch'])
+            if a_row[1]['rv_target'][-2:]=='GG':
+                list_GG.append('OK')
+            else:
+                list_GG.append('NO')  
+        else:
+            if a_row[1]['fw_target'][-2:]=='GG':
+                list_best_target.append(a_row[1]['fw_target'])
+                list_best_location.append(a_row[1]['fw_location'])
+                list_cleavage_site.append(int(a_row[1]['fw_location'].split('-')[1]) - 6)
+                list_delete.append(a_row[1]['fw_deletion'])
+                list_insert.append(a_row[1]['fw_insertion'])
+                list_mismat.append(a_row[1]['fw_mismatch'])
+                list_GG.append('OK_same_score')
+            # 发现没有 GG 则看 RC
+            elif a_row[1]['rv_target'][-2:]=='GG':
+                list_best_target.append(a_row[1]['rv_target'])
+                list_best_location.append(a_row[1]['rv_location'])
+                list_cleavage_site.append(int(a_row[1]['rv_location'].split('-')[0].split(':')[1]) + 5)
+                list_delete.append(a_row[1]['rv_deletion'])
+                list_insert.append(a_row[1]['rv_insertion'])
+                list_mismat.append(a_row[1]['rv_mismatch'])
+                list_GG.append('OK_same_score')
+            else:
+                list_best_target.append(a_row[1]['fw_target'])
+                list_best_location.append(a_row[1]['fw_location'])
+                list_cleavage_site.append(int(a_row[1]['fw_location'].split('-')[1]) - 6)
+                list_delete.append(a_row[1]['fw_deletion'])
+                list_insert.append(a_row[1]['fw_insertion'])
+                list_mismat.append(a_row[1]['fw_mismatch'])                    
+                list_GG.append('NO_same_score')
+    # 记入 df_candidate
+    df_candidate['deletion'] = list_delete
+    df_candidate['insertion'] = list_insert
+    df_candidate['mismatch'] = list_mismat
+    df_candidate['GG'] = list_GG
+    df_candidate['best_target'] = list_best_target
+    df_candidate['target_location'] = list_best_location
+    df_candidate['cleavage_site'] = list_cleavage_site
+    df_candidate = pd.concat([xseq.bedfmt(df_candidate['target_location']), df_candidate], axis=1)
+
+    return df_candidate
+
+def left_realign(dp_bdg_chr, loc_shift_left, ref_fasta, sgRNA_seq, PAM, PAM_loc, n_iter):
+    # print(loc_shift_left)
+    fasta = xseq.get_seq(loc_shift_left, ref_fasta)
+    df_candidate = local_realign(sgRNA_seq, fasta, PAM, PAM_loc)
+    sr_candidate = df_candidate.iloc[0].copy()
+    chrom = sr_candidate['chr']
+    cleavage_site = sr_candidate['cleavage_site']
+    flank_regions = [500]
+    signals = target_signal(dp_bdg_chr.to_pandas(), chrom, cleavage_site, flank_regions=flank_regions)
+    L_neg_1000 = signals[2]
+    # 计算左移后的 L_neg_1000，如果还是负数则迭代，最多迭代 10 次
+    if L_neg_1000 < 0:
+        st = sr_candidate['st']
+        ed = sr_candidate['ed']
+        loc_shift_left = f'{chrom}:{int(st)-1000}-{int(ed)-20}'
+        n_iter += 1
+        if n_iter < 10:
+            return left_realign(dp_bdg_chr, loc_shift_left, ref_fasta, sgRNA_seq, PAM, PAM_loc, n_iter)
+        else:
+            sr_candidate.loc['realign'] = 'fail'
+            return sr_candidate
+    else:
+        sr_candidate.loc['realign'] = 'success'
+        return sr_candidate
+
+def right_realign(dp_bdg_chr, loc_shift_right, ref_fasta, sgRNA_seq, PAM, PAM_loc, n_iter):
+    # print(loc_shift_right)
+    fasta = xseq.get_seq(loc_shift_right, ref_fasta)
+    df_candidate = local_realign(sgRNA_seq, fasta, PAM, PAM_loc)
+    sr_candidate = df_candidate.iloc[0].copy()
+    chrom = sr_candidate['chr']
+    cleavage_site = sr_candidate['cleavage_site']
+    flank_regions = [500]
+    signals = target_signal(dp_bdg_chr.to_pandas(), chrom, cleavage_site, flank_regions=flank_regions)
+    R_neg_1000 = signals[5]
+    # 计算右移后的 R_neg_1000，如果还是负数则迭代，最多迭代 10 次
+    if R_neg_1000 < 0:
+        st = sr_candidate['st']
+        ed = sr_candidate['ed']
+        loc_shift_right = f'{chrom}:{int(st)+20}-{int(ed)+1000}'
+        n_iter += 1
+        if n_iter < 10:
+            return right_realign(dp_bdg_chr, loc_shift_right, ref_fasta, sgRNA_seq, PAM, PAM_loc, n_iter)
+        else:
+            sr_candidate.loc['realign'] = 'fail'
+            return sr_candidate
+    else:
+        sr_candidate.loc['realign'] = 'success'
+        return sr_candidate
+
+

{offtracker-2.12.2 → offtracker-2.13.0}/offtracker/X_sequence.py

@@ -3,7 +3,8 @@ import math
 import pandas as pd
 from itertools import product
 import numpy as np
-import os, glob
+import os, glob, sys
+import polars as pl
 
 ambiguous_nt = {'A': ['A'],
                 'T': ['T'],
@@ -115,7 +116,7 @@ def add_ID(df, chr_col=0, midpoint='cleavage_site'):#, midpoint='midpoint'):
 
 
 
-def detect_fastq(folder, n_subfolder, NGS_type='paired-end'):
+def detect_fastq(folder, n_subfolder, NGS_type='paired-end', skip_trimmed=False):
     """
     搜索 folder 的 n级子目录下的所有 fastq/fastq.gz/fq/fq.gz 文件
     paired-end 模式 : 识别 2.fq/2.fastq 为 paired-end 的 R2 文件，并验证对应 R1 文件
@@ -151,6 +152,9 @@ def detect_fastq(folder, n_subfolder, NGS_type='paired-end'):
             fq_files = glob.glob( os.path.join(folder, n_subfolder*'*/', fastq ) )
             print(f'{len(fq_files)} {fastq[2:]} samples detected')
             files_R2.extend( fq_files )
+
+        if skip_trimmed:
+            files_R2 = [f for f in files_R2 if '_trimmed_2.fq.gz' not in f]
         #
         if len(files_R2) > 0:
             files_R2 = pd.Series(files_R2).sort_values().reset_index(drop=True)
@@ -243,6 +247,137 @@ def sgRNA_alignment(a_key, sgRNA, seq, frag_len, DNA_matrix=None, mismatch_score
     else:
         return [best_alignment.score, position_pct, target, target_location, deletion, insertion, mismatch]
 
+def sgRNA_alignment_new(a_key, sgRNA, seq, substitution_matrix=None, alphabet=None, 
+                   mismatch_score=0.01):
+    """
+    Perform local alignment using Bio.Align instead of deprecated pairwise2.
+    """
+    if substitution_matrix is None or alphabet is None:
+        substitution_matrix, alphabet = create_substitution_matrix(mismatch_score)
+    
+    # Create aligner
+    aligner = Align.PairwiseAligner()
+    aligner.substitution_matrix = Align.substitution_matrices.Array(
+        alphabet=alphabet, dims=2, data=substitution_matrix
+    )
+    aligner.open_gap_score = -2
+    aligner.extend_gap_score = -2
+    aligner.mode = 'local'
+    
+    try:
+        # Perform alignment
+        alignments = aligner.align(sgRNA, seq)
+
+        if not alignments:
+            # No alignment found, return default values
+            return [0, 0, '', f"{a_key.split(':')[0]}:0-0", 0, 0, len(sgRNA)]
+        
+        # Convert to list for indexing
+        alignments = list(alignments)
+         
+        # Extract alignment information
+        coords = alignments[0].coordinates
+        start_target = coords[1][0]
+        end_target = coords[1][-1]
+        
+        # Extract target sequence directly from coordinates
+        # target = seq[start_target:end_target]
+        
+        # Get aligned sequences for detailed analysis
+        alignment_str = str(alignments[0])
+        alignment_lines = alignment_str.split('\n')
+        if len(alignment_lines) >= 3:
+            aligned_sgrna = [x for x in alignment_lines[0].split(' ') if x != '']
+            aligned_genome = [x for x in alignment_lines[2].split(' ') if x != '']
+        else:
+            raise ValueError("Unexpected alignment format")
+        
+        assert int(aligned_sgrna[-1]) == len(sgRNA)
+        
+        # Calculate indels and mismatches
+        # deletion = RNA bulge
+        # insertion = DNA bulge
+        aligned_sgrna_seq = aligned_sgrna[-2]
+        aligned_genome_seq = aligned_genome[-2]
+        insertion = aligned_sgrna_seq.count('-') if '-' in aligned_sgrna_seq else 0
+        deletion = aligned_genome_seq.count('-') if '-' in aligned_genome_seq else 0
+        
+        # Count mismatches by comparing sequences directly
+        # mismatch = 0
+        # assert len(aligned_sgrna_seq) == len(aligned_genome_seq)
+        # for i in range(len(aligned_sgrna_seq)):
+        #     if (aligned_sgrna_seq[i] != aligned_genome_seq[i]) & (aligned_sgrna_seq[i] != 'N') & (aligned_genome_seq[i] != 'N'):
+        #         mismatch += 1
+
+        mismatch = round((alignments[0].score % 1)/mismatch_score)
+        
+        # Calculate target location
+        pos_st = int(a_key.split('-')[0].split(':')[1]) + 1
+        chr_name = a_key.split(':')[0]
+        target_st = pos_st + start_target
+        target_ed = pos_st + end_target - 1
+        target_location = f"{chr_name}:{target_st}-{target_ed}"
+        
+        score = alignments[0].score
+        
+        return [score, aligned_genome_seq, target_location, deletion, insertion, mismatch]
+            
+    except Exception as e:
+        print(f"Alignment error for {a_key}: {e}")
+        return [0, 0, '', f"{a_key.split(':')[0]}:0-0", 0, 0, len(sgRNA)]
+
+
+def get_seq(location, ref_fasta, return_df=False) -> dict:
+    """
+    根据 genome location 取序列
+    location 如果是 list, str 则形式为 "chr1:123456-123458" 或 ["chr1:123456-123458", "chr2:123456-123458"]
+    location 如果是 pd.DataFrame, pl.DataFrame 则默认前三列为 bed 格式
+    ref_fasta 是参考基因组 fasta 文件的路径
+    默认返回字典, key 是位置, value 是序列
+    如果 return_df 为 True, 则返回 pl.DataFrame, 第一列为位置, 第二列为序列
+
+    pybedtools 返回的序列实际上没有包括坐标 start 的那个碱基，这一点和 twoBitToFa 一样
+    但是 IGV/UCSC 等序列是包括 start 的碱基，blast 的结果也是包括 start 的
+    所以在后续分析时要注意这一点
+    """
+    if sys.platform[:3]=='win':
+        # windows 似乎装不了 pybedtools
+        raise ValueError('windows 似乎装不了 pybedtools')
+    else:
+        import pybedtools
+    #########
+    # 根据 genome location 取序列
+    #########
+    if isinstance(location,(list,str)):
+        bed_loc = bedfmt(location)
+    elif isinstance(location,pd.DataFrame):
+        bed_loc = location.iloc[:,:3]
+    elif isinstance(location,pl.DataFrame):
+        bed_loc = location[:,:3]
+    else:
+        raise ValueError('location must be a list, str or pd.DataFrame')
+    
+    fasta = pybedtools.example_filename(ref_fasta)
+    temp_bed = './temp_amp_loc.bed'
+    write_bed(bed_loc, temp_bed)
+    a = pybedtools.BedTool(temp_bed)
+    a = a.sequence(fi=fasta)
+    with open(a.seqfn, encoding='utf-8') as f:
+        dict_seq = {} # 定义一个空的字典
+        for line in f:
+            line = line.strip() # 去除末尾换行符
+            if line[0] == '>':
+                header = line[1:]
+            else:
+                sequence = line
+                dict_seq[header] = dict_seq.get(header,'') + sequence
+    
+    # remove temp_amp_loc.bed
+    os.remove(temp_bed)
+    if return_df:
+        return pl.DataFrame(list(dict_seq.items()), orient='row', schema={'location':pl.String,'sequence':pl.String})
+    else:
+        return dict_seq
 
 def combine_df(list_df, op = 'mean'):
     # df 行列、结构必须一模一样，非数字部分也一模一样，只有数字不同

{offtracker-2.12.2 → offtracker-2.13.0}/offtracker/_version.py

@@ -1,4 +1,4 @@
-__version__ = "2.12.2"
+__version__ = "2.13.0"
 # 2023.08.11. v1.1.0	adding a option for not normalizing the bw file
 # 2023.10.26. v1.9.0	prerelease for v2.0
 # 2023.10.27. v2.0.0	大更新，还没微调
@@ -33,11 +33,13 @@ __version__ = "2.12.2"
 # 2025.04.25. v2.8.0	修复了 offtracker candidates 会把小写序列转换成 N 的 bug
 # 2025.05.22. v2.9.0	翻新部分代码结构
 # 2025.06.05. v2.10.0	增加了QC模块。保留了负数score的记录，并在plot时显示为红字。增加了 "--ignore_chr" 用于跳过common chr过滤。
-# 2025.06.17. v2.10.7   修复翻新代码结构导致的bug
-# 2025.06.27. v2.10.8   将 chmod 放在了 setup.py 里
-# 2025.06.28. v2.10.9   现在 pip 都是从 wheel 安装，不再运行 setup.py，所以增加一个 offtracker_init.py
-# 2025.06.28. v2.10.10  直接塞 script 里试试
-# 2025.06.28. v2.10.11  回滚到2.10.9外加修正
-# 2025.07.02. v2.11.4   基于 blast 的缺陷更新 candidates，去除 quick mode
-# 2025.07.04. v2.11.5   offtracker_analysis 提前 skip 已有结果的样本
-# 2025.07.04. v2.12.2   新增 region_index 标记区域，用于更好的去重
+# 2025.06.17. v2.10.7	修复翻新代码结构导致的bug
+# 2025.06.27. v2.10.8	将 chmod 放在了 setup.py 里
+# 2025.06.28. v2.10.9	现在 pip 都是从 wheel 安装，不再运行 setup.py，所以增加一个 offtracker_init.py
+# 2025.06.28. v2.10.10	直接塞 script 里试试
+# 2025.06.28. v2.10.11	回滚到2.10.9外加修正
+# 2025.07.02. v2.11.4	基于 blast 的缺陷更新 candidates，去除 quick mode
+# 2025.07.04. v2.11.5	offtracker_analysis 提前 skip 已有结果的样本
+# 2025.07.04. v2.12.2	新增 region_index 标记区域，用于更好的去重
+# 2025.07.18. v2.12.3	新增QC自动避免重复读取 trimmed fastq files
+# 2025.08.08. v2.13.0	测试 local realign 功能

{offtracker-2.12.2 → offtracker-2.13.0/offtracker.egg-info}/PKG-INFO

@@ -1,6 +1,6 @@
 Metadata-Version: 2.1
 Name: offtracker
-Version: 2.12.2
+Version: 2.13.0
 Summary: Tracking-seq data analysis
 Home-page: https://github.com/Lan-lab/offtracker
 Author: Runda Xu

{offtracker-2.12.2 → offtracker-2.13.0}/scripts/offtracker_analysis.py

@@ -83,7 +83,7 @@ def main():
         df_candidate.index = df_candidate['target_location']
         df_candidate_brief = df_candidate[['chr','st','ed','best_strand','best_target','best_seq_score',
                                  'deletion', 'insertion','mismatch', 'GG', 
-                                 'target_location', 'cleavage_site', 'ID_1','ID_2', 'region_index']] # 2025.07.06 添加 region_index
+                                 'target_location', 'cleavage_site', 'region_index']] # 2025.07.06 添加 region_index, 去除 'ID_1','ID_2',
         df_candidate_sub = df_candidate[['chr','cleavage_site']]
     except FileNotFoundError:
         return 'Please run offtracker_candidates.py first and provide the correct directory with --seqfolder'
@@ -366,9 +366,9 @@ def main():
     # 2024.06.03. 以防 fdr<=fdr_thresh 滤掉了 track_score>=2 的位点
     bool_fdr = df_result['fdr']<=fdr_thresh
     bool_score = df_result['track_score']>=score_thresh
-    # 2025.06.05. BE可能会形成单边信号，导致 track_score 为负数，也保留
-    bool_neg_score = df_result['track_score']< -1
-    df_output = df_result[bool_fdr|bool_score|bool_neg_score].copy()
+    # 2025.06.05. BE可能会形成单边信号，但是很少见，如果 control 用的是别的 sgRNA 的样本，对应脱靶位置附近一般就是负数
+    # bool_neg_score = df_result['track_score']< -1
+    df_output = df_result[bool_fdr|bool_score].copy()
     if pattern_ctr != 'none':
         df_output = df_output[['target_location', 'best_strand','best_target','deletion','insertion','mismatch',
                             'exp_L_length', 'exp_R_length','ctr_L_length','ctr_R_length','L_length','R_length','signal_length',
@@ -383,6 +383,13 @@ def main():
         df_output.columns = ['target_location', 'strand', 'target', 'deletion', 'insertion', 'mismatch', 
                             'L_length', 'R_length','signal_length',
                             'seq_score', 'track_score', 'log2_track_score','FDR', 'rank']
+    
+
+    # 2025.08.08. 增加对阳性位点的 target_location 重比对功能，避免 blast 比对后的 realign 在更大范围内的存在不准确的情况
+
+
+
+    
     df_output.to_csv(f'Offtracker_result_{outname}.csv', index=False)
 
     if args.clean:

{offtracker-2.12.2 → offtracker-2.13.0}/scripts/offtracker_qc.py

@@ -29,7 +29,8 @@ def main():
     parser.add_argument('-o','--outdir', type=str, default='same',       help='The output folder')
     parser.add_argument('--subfolder'  , type=int, default=0,            help='subfolder level')
     parser.add_argument('-t','--thread', type=int, default=8,            help='Number of threads to be used')
-
+    parser.add_argument('--include_trimmed', action='store_true',        help='Do not skip trimmed fastq files')
+    
     args = parser.parse_args()
 
     # 自动化的参数调整和报错
@@ -42,7 +43,10 @@ def main():
             os.makedirs(args.outdir)
 
     # 搜索 folder 的 n级子目录下的所有 fastq/fastq.gz/fq/fq.gz 文件
-    sample_names, files_R1, files_R2 = xseq.detect_fastq(args.folder, n_subfolder=args.subfolder)
+    if args.include_trimmed:
+        sample_names, files_R1, files_R2 = xseq.detect_fastq(args.folder, n_subfolder=args.subfolder)
+    else:
+        sample_names, files_R1, files_R2 = xseq.detect_fastq(args.folder, n_subfolder=args.subfolder, skip_trimmed=True)
 
     assert not isinstance(sample_names, str), 'No fastq file is detected!'