ORForise - 原核生物基因组注释分析和比较平台

发表于生物信息学： https ://academic.oup.com/bioinformatics/article/38/5/1198/6454948

用于分析和比较原核生物编码序列 (CDS) 基因预测的平台。

可以将新的基因组注释与来自 Ensembl 的提供的参考注释和来自其他工具（或任何给定的 GFF 注释）的预测进行比较。

要求和安装：

ORForise 平台是用 Python3.8 编写的，只需要 NumPy 库（安装 ORForise 时应由 pip 自动安装），这是大多数 Python3 基础安装的标准。

安装：

ORForise 平台可通过 pip Python 包管理器获得pip3 install ORForise。

考虑将“--no-cache-dir”与 pip 一起使用，以确保下载最新版本的软件包。

所需文件：

要运行，您需要：

• 输入基因组 FASTA 和相应的 GFF 文件（或带有注释基因的 CDS 预测，您想在下面列出的工具输出格式之一中用作参考）。
• 来自同一基因组的兼容工具之一的预测输出。

如何添加自己的基因组：

必须为要进行分析的基因组提供相应的 FASTA 和 GFF 文件，包括任何要比较的工具的相应输出。

如何添加自己的工具：

如果新工具在 GFF 中报告其预测，您可以使用“GFF”为 ORForise 提供参考-rt或预测-t选项。如果该工具使用另一种非标准格式，则可以通过 GitHub 请求将其添加为选项。

测试：

~ORForise/TestingGitHub 存储库的目录中提供了预先计算的测试和数据，其中包括下面介绍的所有工具的示例输入和输出文件。和Annotation-Compare的示例输出文件可用于验证安装。GFF-AdderGFF-Intersector

CDS预测分析：

用例：（如果通过 pip 运行）

求助：Annotation-Compare -h

usage: Annotation-Compare [-h] -dna GENOME_DNA [-rt REFERENCE_TOOL] -ref
                             REFERENCE_ANNOTATION -t TOOL -tp TOOL_PREDICTION
                             [-o OUTNAME] [-v {True,False}]

optional arguments:
  -h, --help            show this help message and exit
  -dna GENOME_DNA, --genome_DNA GENOME_DNA
                        Genome DNA file (.fa) which both annotations are based
                        on
  -rt REFERENCE_TOOL, --reference_tool REFERENCE_TOOL
                        What type of Annotation to compare to? -- Leave blank
                        for Ensembl reference- Provide tool name to compare
                        output from two tools (GeneMarkS)
  -ref REFERENCE_ANNOTATION, --reference_annotation REFERENCE_ANNOTATION
                        Which reference annotation file to use as reference?
  -t TOOL, --tool TOOL  Which tool to analyse? (Prodigal)
  -tp TOOL_PREDICTION, --tool_prediction TOOL_PREDICTION
                        Tool genome prediction file (.gff) - Different Tool
                        Parameters are compared individually via separate
                        files
  -o OUTNAME, --outname OUTNAME
                        Define full output filename (format is CSV) - If not
                        provided, summary will be printed to std-out
  -v {True,False}, --verbose {True,False}
                        Default - False: Print out runtime status

将新的基因组注释与 Ensembl 注释进行比较：

基因组注释是一个困难的过程，即使对于原核生物也是如此。ORForise 允许直接和系统地分析从各种工具到参考基因组注释的新型 CDS 预测，例如由 Ensembl Bacteria 提供的那些。

示例：通过 pip 安装将允许用户直接从 ORForise 包中调用程序。

Annotation-Compare -dna ~/Testing/Myco.fa -ref ~/Testing/Myco.gff -t Prodigal -tp ~/Testing/Prodigal_Myco.gff

在单个基因组上相互比较不同的新注释：

如果参考基因组注释不可用或需要两个或更多工具之间的直接比较，可以使用 ORForise 作为下面的示例。

聚合 CDS 预测分析：

用例：（如果通过 pip 运行）

求助：Aggregate-Compare -h

usage: Aggregate-Compare [-h] -dna GENOME_DNA -t TOOLS -tp TOOL_PREDICTIONS
                            [-rt REFERENCE_TOOL] -ref REFERENCE_ANNOTATION
                            [-o OUTNAME] [-v {True,False}]

optional arguments:
  -h, --help            show this help message and exit
  -dna GENOME_DNA, --genome_DNA GENOME_DNA
                        Genome DNA file (.fa) which both annotations are based
                        on
  -t TOOLS, --tools TOOLS
                        Which tools to analyse? (Prodigal,GeneMarkS)
  -tp TOOL_PREDICTIONS, --tool_predictions TOOL_PREDICTIONS
                        Tool genome prediction file (.gff) - Providefile
                        locations for each tool comma separated
  -rt REFERENCE_TOOL, --reference_tool REFERENCE_TOOL
                        What type of Annotation to compare to? -- Leave blank
                        for Ensembl reference- Provide tool name to compare
                        output from two tools (GeneMarkS)
  -ref REFERENCE_ANNOTATION, --reference_annotation REFERENCE_ANNOTATION
                        Which reference annotation file to use as reference?
  -o OUTNAME, --outname OUTNAME
                        Define full output filename (format is CSV) - If not
                        provided, summary will be printed to std-out
  -v {True,False}, --verbose {True,False}
                        Default - False: Print out runtime status

例子：

Aggregate-Compare -ref ~/Testing/Myco.gff -dna ~/Testing/Myco.fa -t Prodigal,TransDecoder,GeneMark_S_2 -tp ~/Testing/Prodigal_Myco.gff,~/Testing/TransDecoder_Myco.gff,~/Testing/GeneMark_S_2_Myco.gff

这将把 Prodigal、TransDecoder 和 GLIMMER 3 的聚合预测与 Ensembl Bacteria 提供的支原体参考注释进行比较。

注释比较输出 - Annotation_Compare 和 Aggregate_Compare 的输出格式相同：

打印到屏幕示例 - Prodigal 预测与大肠杆菌的 Ensembl Bacteria 参考注释相比：

Annotation-Compare.py  -ref ./Testing/Myco.gff -dna ./Testing/Myco.fa -t Prodigal -tp ./Testing/Prodigal_Myco.gff
Genome Used: Myco
Reference Used: Testing/Myco.gff
Tool Compared: Prodigal
Perfect Matches:128[476] -26.89%
Partial Matches:62[476] - 13.03%
Missed Genes:286[476] - 60.08%
Complete

Aggregate-Compare -ref ./Testing/Myco.gff -dna ./Testing/Myco.fa -t Prodigal,TransDecoder,GeneMark_S_2 -tp ./Testing/Prodigal_Myco.gff,./Testing/TransDecoder_Myco.gff,./Testing/GeneMark_S_2_Myco.gff
Prodigal
TransDecoder
GeneMark_S_2
Match filtered out
Match filtered out
Match filtered out
Match filtered out
Match filtered out
Match filtered out
Genome Used: Myco
Reference Used: ./Testing/Myco.gff
Tools Compared: Prodigal,TransDecoder,GeneMark_S_2
Perfect Matches:132[476]
Partial Matches:58[476]
Missed Genes:286[476]

这是比较工具的默认输出。

'-o' 输出到 CSV 文件的示例 - 与大肠杆菌的 Ensembl Bacteria 参考注释相比的败家预测：

输出设计为人类可读且可通过包含的“ORForise_Analysis”脚本进行解释。下面的示例展示了 12 个“代表性”和 72 个“所有”指标，但仅显示了每个单独预测报告的一个条目（Perfect_Match_Genes、Partial_Match_Genes、Missed_Genes、Predicted_CDS_Without_Corresponding_Gene_in_Reference、Predicted_CDSs_Which_Detected_more_than_one_Gene）。

Representative_Metrics:
Percentage_of_Genes_Detected,Percentage_of_ORFs_that_Detected_a_Gene,Percent_Difference_of_All_ORFs,Median_Length_Difference,Percentage_of_Perfect_Matches,Median_Start_Difference_of_Matched_ORFs,Median_Stop_Difference_of_Matched_ORFs,Percentage_Difference_of_Matched_Overlapping_CDSs,Percent_Difference_of_Short-Matched-ORFs,Precision,Recall,False_Discovery_Rate
39.92,19.10,109.03,-62.17,67.37,67.5,-85.5,-83.71,-17.39,0.19,0.40,0.81
All_Metrics:
Number_of_ORFs,Percent_Difference_of_All_ORFs,Number_of_ORFs_that_Detected_a_Gene,Percentage_of_ORFs_that_Detected_a_Gene,Number_of_Genes_Detected,Percentage_of_Genes_Detected,Median_Length_of_All_ORFs,Median_Length_Difference,Minimum_Length_of_All_ORFs,Minimum_Length_Difference,Maximum_Length_of_All_ORFs,Maximum_Length_Difference,Median_GC_content_of_All_ORFs,Percent_Difference_of_All_ORFs_Median_GC,Median_GC_content_of_Matched_ORFs,Percent_Difference_of_Matched_ORF_GC,Number_of_ORFs_which_Overlap_Another_ORF,Percent_Difference_of_Overlapping_ORFs,Maximum_ORF_Overlap,Median_ORF_Overlap,Number_of_Matched_ORFs_Overlapping_Another_ORF,Percentage_Difference_of_Matched_Overlapping_CDSs,Maximum_Matched_ORF_Overlap,Median_Matched_ORF_Overlap,Number_of_Short-ORFs,Percent_Difference_of_Short-ORFs,Number_of_Short-Matched-ORFs,Percent_Difference_of_Short-Matched-ORFs,Number_of_Perfect_Matches,Percentage_of_Perfect_Matches,Number_of_Perfect_Starts,Percentage_of_Perfect_Starts,Number_of_Perfect_Stops,Percentage_of_Perfect_Stops,Number_of_Out_of_Frame_ORFs,Number_of_Matched_ORFs_Extending_a_Coding_Region,Percentage_of_Matched_ORFs_Extending_a_Coding_Region,Number_of_Matched_ORFs_Extending_Start_Region,Percentage_of_Matched_ORFs_Extending_Start_Region,Number_of_Matched_ORFs_Extending_Stop_Region,Percentage_of_Matched_ORFs_Extending_Stop_Region,Number_of_All_ORFs_on_Positive_Strand,Percentage_of_All_ORFs_on_Positive_Strand,Number_of_All_ORFs_on_Negative_Strand,Percentage_of_All_ORFs_on_Negative_Strand,Median_Start_Difference_of_Matched_ORFs,Median_Stop_Difference_of_Matched_ORFs,ATG_Start_Percentage,GTG_Start_Percentage,TTG_Start_Percentage,ATT_Start_Percentage,CTG_Start_Percentage,Other_Start_Codon_Percentage,TAG_Stop_Percentage,TAA_Stop_Percentage,TGA_Stop_Percentage,Other_Stop_Codon_Percentage,True_Positive,False_Positive,False_Negative,Precision,Recall,False_Discovery_Rate,Nucleotide_True_Positive,Nucleotide_False_Positive,Nucleotide_True_Negative,Nucleotide_False_Negative,Nucleotide_Precision,Nucleotide_Recall,Nucleotide_False_Discovery_Rate,ORF_Nucleotide_Coverage_of_Genome,Matched_ORF_Nucleotide_Coverage_of_Genome
995,109.03,190,19.10,190,39.92,335.0,-62.17,89,-21.24,3152,-41.81,31.50,0.20,32.83,4.42,279,26.24,135,0.00,36,-83.71,31,4.50,443,1826.09,19,-17.39,128,67.37,162,85.26,154,81.05,0,0,0.00,4,2.11,0,0.00,570,0.57,425,0.43,67.5,-85.5,63.12,15.28,21.61,0.00,0.00,0.00,11.06,27.44,61.51,0.00,0.40,1.69,0.60,0.19,0.40,0.81,0.82,0.31,0.69,0.18,0.96,0.82,0.04,77.15,24.47
CDS_Gene_Coverage_of_Genome:
90.62
Start_Position_Difference:
-78,33,93,294,144,408,3,18,156,-42,45,90,333,333,-39,111,201,93,120,-354,-150,-366,117,-138,-240,123,-153,-51
Stop_Position_Difference:
-192,-147,108,-216,87,-678,-96,-156,-321,-240,-168,-162,-51,-126,-33,-3,-93,-12,-204,-189,-156,237,-45,-219,-201,-537,-30,-78,159,243,60,21,15,183,288,6
Alternative_Starts_Predicted:

Alternative_Stops_Predicted:

Undetected_Gene_Metrics:
ATG_Start ,GTG_Start ,TTG_Start ,ATT_Start ,CTG_Start ,Alternative_Start_Codon ,TGA_Stop ,TAA_Stop ,TAG_Stop ,Alternative_Stop_Codon ,Median_Length ,ORFs_on_Positive_Strand ,ORFs_on_Negative_Strand
88.46,7.69,3.85,0.00,0.00,0.00,0.00,74.13,25.87,0.00,1047.50,156,130
Perfect_Match_Genes:
>Myco_686_1828_+ 
ATGAAAATATTAATTAATAAAAGTGAATTGAATAAAATTTTGAAAAAAATGAATAACGTTATTATTTCCAATAACAAAATAAAACCACATCATTCATATTTTTTAATAGAGGCAAAAGAAAAAGAAATAAACTTTTATGCTAACAATGAATACTTTTCTGTCAAATGTAATTTAAATAAAAATATTGATATTCTTGAACAAGGCTCCTTAATTGTTAAAGGAAAAATTTTTAACGATCTTATTAATGGCATAAAAGAAGAGATTATTACTATTCAAGAAAAAGATCAAACACTTTTGGTTAAAACAAAAAAAACAAGTATTAATTTAAACACAATTAATGTGAATGAATTTCCAAGAATAAGGTTTAATGAAAAAAACGATTTAAGTGAATTTAATCAATTCAAAATAAATTATTCACTTTTAGTAAAAGGCATTAAAAAAATTTTTCACTCAGTTTCAAATAATCGTGAAATATCTTCTAAATTTAATGGAGTAAATTTCAATGGATCCAATGGAAAAGAAATATTTTTAGAAGCTTCTGACACTTATAAACTATCTGTTTTTGAGATAAAGCAAGAAACAGAACCATTTGATTTCATTTTGGAGAGTAATTTACTTAGTTTCATTAATTCTTTTAATCCTGAAGAAGATAAATCTATTGTTTTTTATTACAGAAAAGATAATAAAGATAGCTTTAGTACAGAAATGTTGATTTCAATGGATAACTTTATGATTAGTTACACATCGGTTAATGAAAAATTTCCAGAGGTAAACTACTTTTTTGAATTTGAACCTGAAACTAAAATAGTTGTTCAAAAAAATGAATTAAAAGATGCACTTCAAAGAATTCAAACTTTGGCTCAAAATGAAAGAACTTTTTTATGCGATATGCAAATTAACAGTTCTGAATTAAAAATAAGAGCTATTGTTAATAATATCGGAAATTCTCTTGAGGAAATTTCTTGTCTTAAATTTGAAGGTTATAAACTTAATATTTCTTTTAACCCAAGTTCTCTATTAGATCACATAGAGTCTTTTGAATCAAATGAAATAAATTTTGATTTCCAAGGAAATAGTAAGTATTTTTTGATAACCTCTAAAAGTGAACCTGAACTTAAGCAAATATTGGTTCCTTCAAGATAA 

>Myco_4812_7322_+ 
ATGGCAAAGCAACAAGATCAAGTAGATAAGATTCGTGAAAACTTAGACAATTCAACTGTCAAAAGTATTTCATTAGCAAATGAACTTGAGCGTTCATTCATGGAATATGCTATGTCAGTTATTGTTGCTCGTGCTTTACCTGATGCTAGAGATGGACTTAAACCAGTTCATCGTCGTGTTCTTTATGGTGCTTATATTGGTGGCATGCACCATGATCGTCCTTTTAAAAAGTCTGCGAGGATTGTTGGTGATGTAATGAGTAAATTCCACCCTCATGGTGATATGGCAATATATGACACCATGTCAAGAATGGCTCAAGACTTTTCATTAAGATACCTTTTAATTGATGGTCATGGTAATTTTGGTTCTATAGATGGTGATAGACCTGCTGCACAACGTTATACAGAAGCAAGATTATCTAAACTTGCAGCAGAACTTTTAAAAGATATTGATAAAGATACAGTTGACTTTATTGCTAATTATGATGGTGAGGAAAAAGAACCAACTGTTCTACCAGCAGCTTTCCCTAACTTACTTGCAAATGGTTCTAGTGGGATTGCAGTTGGAATGTCAACATCTATTCCTTCCCATAATCTCTCTGAATTAATTGCGGGTTTAATCATGTTAATTGATAATCCTCAATGCACTTTTCAAGAATTATTAACTGTAATTAAAGGACCTGATTTTCCAACAGGAGCTAACATTATCTACACAAAAGGAATTGAAAGCTACTTTGAAACAGGTAAAGGCAATGTAGTAATTCGTTCTAAAGTTGAGATAGAACAATTGCAAACAAGAAGTGCATTAGTTGTAACTGAAATTCCTTACATGGTTAACAAAACTACCTTAATTGAAAAGATTGTAGAACTTGTTAAAGCTGAAGAGATTTCAGGAATTGCTGATATCCGTGATGAATCCTCTCGAGAAGGAATAAGGTTAGTGATTGAAGTAAAACGCGACACTGTACCTGAAGTTTTATTAAATCAACTTTTTAAATCAACAAGATTACAAGTACGCTTCCCTGTTAATATGCTTGCTTTAGTTAAAGGAGCTCCTGTACTTCTCAACATGAAACAAGCTTTGGAAGTATATCTTGATCATCAAATTGATGTTCTTGTTAGAAAAACAAAGTTTGTGCTTAATAAACAACAAGAACGTTATCACATTTTAAGCGGACTTTTAATTGCTGCTTTAAATATTGATGAGGTTGTTGCAATTATTAAAAAATCAGCAAATAACCAGGAAGCAATTAATACATTAAATACAAAGTTTAAGCTTGATGAAATTCAAGCTAAAGCAGTTCTTGACATGCGTTTAAGGAGCTTAAGCGTACTTGAAGTTAACAAACTTCAAACTGAACAAAAAGAGTTAAAAGATTCAATTGAATTTTGTAAGAAAGTGTTAGCTGATCAAAAATTACAGCTAAAAATAATCAAAGAGGAATTGCAAAAAATCAATGATCAGTTTGGTGATGAAAGAAGAAGTGAAATTCTCTATGATATCTCTGAGGAAATTGATGATGAATCATTGATAAAAGTTGAGAATGTAGTGATAACTATGTCTACAAATGGTTATCTAAAAAGGATTGGAGTTGATGCTTATAATCTTCAACATCGTGGTGGAGTTGGGGTTAAAGGGCTAACTACTTATGTTGATGATAGTATTAGTCAATTATTGGTCTGTTCAACTCACTCTGACTTATTATTTTTTACTGATAAGGGTAAGGTTTATAGAATTAGAGCTCATCAAATTCCCTATGGTTTTAGAACAAATAAAGGTATTCCCGCTGTTAACTTAATCAAAATTGAAAAGGATGAAAGAATTTGTTCATTGTTATCTGTTAATAACTATGATGATGGTTATTTCTTTTTCTGTACTAAAAATGGAATTGTTAAAAGAACGAGCTTGAATGAATTCATCAACATCTTAAGTAATGGTAAGCGGGCTATATCTTTTGATGATAATGACACTTTGTATTCAGTAATTAAAACCCACGGAAATGATGAGATTTTTATTGGTTCTACCAATGGATTTGTTGTTCGCTTCCATGAAAATCAACTCAGAGTTCTTTCAAGAACAGCAAGAGGTGTATTTGGTATCAGTTTAAATAAAGGAGAATTTGTTAATGGACTATCAACTTCAAGCAACGGTAGCTTACTTTTATCAGTCGGTCAAAATGGAATAGGTAAATTAACGAGCATAGATAAATATAGACTCACAAAACGTAATGCTAAGGGAGTTAAAACTCTAAGGGTTACTGATAGAACAGGCCCTGTTGTTACAACAACCACTGTTTTTGGTAATGAGGATCTTTTAATGATTTCCTCTGCTGGTAAAATTGTGCGTACCAGTTTACAAGAACTTTCAGAACAAGGTAAAAACACTTCTGGTGTTAAGTTAATTAGATTAAAAGATAATGAACGTTTAGAAAGAGTAACTATCTTTAAAGAAGAGTTAGAAGACAAAGAAATGCAACTAGAAGATGTTGGATCCAAACAAATTACGCAATAA 
.........
Partial_Match_Genes:
Gene:9923_11251_+_ATG_TAA 
ATGAAAAGCGAAATTAATATTTTTGCACTAGCAACTGCACCTTTTAATAGTGCATTACATATTATTAGGTTTTCTGGTCCTGATGTTTATGAGATTTTAAACAAGATAACTAATAAAAAAATAACAAGAAAAGGGATGCAAATTCAACGCACATGGATAGTTGATGAAAACAATAAGCGAATTGATGATGTGCTATTATTTAAATTTGTCTCTCCAAATTCTTATACAGGAGAAGATTTAATTGAAATTTCTTGTCATGGTAACATGTTGATCGTTAATGAAATTTGCGCACTTCTTTTAAAAAAAGGAGGTGTTTATGCCAAACCTGGTGAATTTACCCAAAGGAGTTTTTTAAATGGAAAAATGAGTTTACAACAAGCTAGTGCTGTAAATAAATTGATTTTATCTCCTAACTTATTAGTTAAAGATATAGTCTTAAATAATTTAGCGGGTGAAATGGATCAACAATTAGAACAAATAGCTCAACAAGTTAATCAATTAGTAATGCAAATGGAAGTAAACATTGATTATCCAGAATATCTTGATGAACAAGTAGAACTATCAACTTTAAATAATAAAGTTAAATTGATTATTGAAAAGCTTAAAAGAATTATTGAAAATAGTAAACAACTCAAAAAACTTCACGATCCTTTTAAAATTGCCATTATAGGCGAAACTAATGTAGGTAAATCTTCTTTACTCAACGCTTTATTAAATCAAGATAAAGCGATAGTTTCAAATATTAAAGGTAGTACACGCGATGTTGTTGAAGGGGATTTCAATTTAAATGGTTATTTAATCAAGATCTTAGATACTGCAGGTATCCGTAAACATAAAAGTGGGCTTGAAAAAGCAGGAATTAAAAAAAGCTTTGAATCTATAAAGCAAGCTAATTTGGTTATTTATCTTTTAGATGCAACACATCCAAAGAAAGATCTTGAATTAATTAGTTTTTTTAAGAAAAATAAAAAGGATTTTTTTGTTTTCTATAACAAAAAAGATTTAATTACAAATAAGTTTGAAAATAGTATTTCTGCAAAGCAAAAAGATATTAAAGAATTAGTTGATTTATTAACTAAATATATTAACGAGTTTTATAAAAAAATAGATCAAAAAATCTATCTGATTGAAAATTGACAGCAAATTTTAATTGAAAAAATTAAAGAACAATTAGAACAGTTTTTAAAGCAACAAAAAAAATATTTATTTTTCGATGTTTTAGTTACCCATCTAAGAGAAGCTCAACAAGATATTCTTAAACTACTAGGTAAGGATGTAGGTTTTGATTTAGTTAATGAAATTTTTAATAATTTTTGTTTAGGAAAATAA 
ORF:9923_11059_+_ATG_TGA 
ATGAAAAGCGAAATTAATATTTTTGCACTAGCAACTGCACCTTTTAATAGTGCATTACATATTATTAGGTTTTCTGGTCCTGATGTTTATGAGATTTTAAACAAGATAACTAATAAAAAAATAACAAGAAAAGGGATGCAAATTCAACGCACATGGATAGTTGATGAAAACAATAAGCGAATTGATGATGTGCTATTATTTAAATTTGTCTCTCCAAATTCTTATACAGGAGAAGATTTAATTGAAATTTCTTGTCATGGTAACATGTTGATCGTTAATGAAATTTGCGCACTTCTTTTAAAAAAAGGAGGTGTTTATGCCAAACCTGGTGAATTTACCCAAAGGAGTTTTTTAAATGGAAAAATGAGTTTACAACAAGCTAGTGCTGTAAATAAATTGATTTTATCTCCTAACTTATTAGTTAAAGATATAGTCTTAAATAATTTAGCGGGTGAAATGGATCAACAATTAGAACAAATAGCTCAACAAGTTAATCAATTAGTAATGCAAATGGAAGTAAACATTGATTATCCAGAATATCTTGATGAACAAGTAGAACTATCAACTTTAAATAATAAAGTTAAATTGATTATTGAAAAGCTTAAAAGAATTATTGAAAATAGTAAACAACTCAAAAAACTTCACGATCCTTTTAAAATTGCCATTATAGGCGAAACTAATGTAGGTAAATCTTCTTTACTCAACGCTTTATTAAATCAAGATAAAGCGATAGTTTCAAATATTAAAGGTAGTACACGCGATGTTGTTGAAGGGGATTTCAATTTAAATGGTTATTTAATCAAGATCTTAGATACTGCAGGTATCCGTAAACATAAAAGTGGGCTTGAAAAAGCAGGAATTAAAAAAAGCTTTGAATCTATAAAGCAAGCTAATTTGGTTATTTATCTTTTAGATGCAACACATCCAAAGAAAGATCTTGAATTAATTAGTTTTTTTAAGAAAAATAAAAAGGATTTTTTTGTTTTCTATAACAAAAAAGATTTAATTACAAATAAGTTTGAAAATAGTATTTCTGCAAAGCAAAAAGATATTAAAGAATTAGTTGATTTATTAACTAAATATATTAACGAGTTTTATAAAAAAATAGATCAAAAAATCTATCTGATTGAAAATTGA 

Gene:11251_12039_+_ATG_TAA 
ATGGAATACTTTGATGCACATTGTCATTTAAATTGTGAACCTTTACTGAGTGAAATTGAAAAAAGCATCGCTAATTTCAAATTAATTAATTTAAAAGCAAATGTTGTAGGTACAGATTTGGATAATTCTAAAATTGCTGTTGAATTAGCTAAAAAATATCCTGATCTTTTAAAAGCAACCATAGGTATCCATCCAAATGATGTTCATTTAGTTGATTTTAAAAAGACAAAAAAACAACTTAATGAACTATTAATAAATAACAGAAATTTCATAAGTTGTATTGGTGAATATGGTTTTGATTATCACTACACAACAGAATTTATTGAATTGCAAAACAAATTCTTTGAGATGCAATTTGAAATAGCTGAAACTAATAAATTGGTTCACATGCTTCATATTCGTGATGCTCATGAAAAAATTTATGAAATATTAACAAGATTAAAGCCAACTCAACCTGTGATTTTTCATTGTTTCAGTCAAGATATAAATATTGCTAAAAAGCTACTATCATTAAAAGATTTAAATATTGACATCTTCTTTTCTATCCCAGGGATAGTTACTTTTAAGAATGCTCAAGCATTACATGAAGCTTTAAAGATTATTCCTAGTGAATTACTTTTAAGTGAAACTGACTCACCGTGATTAACCCCTTCTCCTTTTCGAGGCAAAGTTAACTGACCTGAATATGTAGTTCATACTGTTAGCACTGTTGCTGAAATAAAAAAAATAGAAATTGCTGAAATGAAGCGAATTATTGTTAAAAATGCAAAAAAATTATTTTGACATTAA 
ORF:11251_11892_+_ATG_TGA 
ATGGAATACTTTGATGCACATTGTCATTTAAATTGTGAACCTTTACTGAGTGAAATTGAAAAAAGCATCGCTAATTTCAAATTAATTAATTTAAAAGCAAATGTTGTAGGTACAGATTTGGATAATTCTAAAATTGCTGTTGAATTAGCTAAAAAATATCCTGATCTTTTAAAAGCAACCATAGGTATCCATCCAAATGATGTTCATTTAGTTGATTTTAAAAAGACAAAAAAACAACTTAATGAACTATTAATAAATAACAGAAATTTCATAAGTTGTATTGGTGAATATGGTTTTGATTATCACTACACAACAGAATTTATTGAATTGCAAAACAAATTCTTTGAGATGCAATTTGAAATAGCTGAAACTAATAAATTGGTTCACATGCTTCATATTCGTGATGCTCATGAAAAAATTTATGAAATATTAACAAGATTAAAGCCAACTCAACCTGTGATTTTTCATTGTTTCAGTCAAGATATAAATATTGCTAAAAAGCTACTATCATTAAAAGATTTAAATATTGACATCTTCTTTTCTATCCCAGGGATAGTTACTTTTAAGAATGCTCAAGCATTACATGAAGCTTTAAAGATTATTCCTAGTGAATTACTTTTAAGTGAAACTGACTCACCGTGA 
.......
Missed_Genes:
>Myco_1828_2760_+ 
ATGAATCTTTACGATCTTTTAGAACTACCAACTACAGCATCAATAAAAGAAATAAAAATTGCTTATAAAAGATTAGCAAAGCGTTATCACCCTGATGTAAATAAATTAGGTTCGCAAACTTTTGTTGAAATTAATAATGCTTATTCAATATTAAGTGATCCTAACCAAAAGGAAAAATATGATTCAATGCTGAAAGTTAATGATTTTCAAAATCGCATCAAAAATTTAGATATTAGTGTTAGATGACATGAAAATTTCATGGAAGAACTCGAACTTCGTAAGAACTGAGAATTTGATTTTTTTTCATCTGATGAAGATTTCTTTTATTCTCCATTTACAAAAAACAAATATGCTTCCTTTTTAGATAAAGATGTTTCTTTAGCTTTTTTTCAGCTTTACAGCAAGGGCAAAATAGATCATCAATTGGAAAAATCTTTATTGAAAAGAAGAGATGTAAAAGAAGCTTGTCAACAGAATAAAAATTTTATTGAAGTTATAAAAGAGCAATATAACTATTTTGGTTGAATTGAAGCTAAGCGTTATTTCAATATTAATGTTGAACTTGAGCTCACACAGAGAGAGATAAGAGATAGAGATGTTGTTAACCTACCTTTAAAAATTAAAGTTATTAATAATGATTTTCCAAATCAACTCTGATATGAAATTTATAAAAACTATTCATTTCGCTTATCTTGAGATATAAAAAATGGTGAAATTGCTGAATTTTTCAATAAAGGTAATAGAGCTTTAGGATGAAAAGGTGACTTAATTGTCAGAATGAAAGTAGTTAATAAAGTAAACAAAAGACTGCGTATTTTTTCAAGCTTTTTTGAGAACGATAAATCTAAATTATGGTTCCTTGTTCCAAACGATAAACAAAGTAATCCTAATAAGGGCGTTTTTAACTATAAAACTCAGCACTTTATTGATTAA 

>Myco_2845_4797_+ 
ATGGAAGAAAATAACAAAGCAAATATCTATGACTCTAGTAGCATTAAGGTCCTTGAAGGACTTGAGGCTGTTAGAAAACGCCCTGGAATGTACATTGGTTCTACTGGCGAAGAAGGTTTGCATCACATGATCTGAGAGATAGTAGACAACTCAATTGATGAAGCAATGGGAGGTTTTGCCAGTTTTGTTAAGCTTACCCTTGAAGATAATTTTGTTACCCGTGTAGAGGATGATGGAAGAGGGATACCTGTTGATATCCATCCTAAGACTAATCGTTCTACAGTTGAAACAGTTTTTACAGTTCTACACGCTGGCGGTAAATTTGATAACGATAGCTATAAAGTGTCAGGTGGTTTACACGGTGTTGGTGCATCAGTTGTTAATGCGCTTAGTTCTTCTTTTAAAGTTTGAGTTTTTCGTCAAAATAAAAAGTATTTTCTCAGCTTTAGCGATGGAGGAAAGGTAATTGGAGATTTGGTCCAAGAAGGTAACTCTGAAAAAGAGCATGGAACAATTGTTGAGTTTGTTCCTGATTTCTCTGTAATGGAAAAGAGTGATTACAAACAAACTGTAATTGTAAGCAGACTCCAGCAATTAGCTTTTTTAAACAAGGGAATAAGAATTGACTTTGTTGATAATCGTAAACAAAACCCACAGTCTTTTTCTTGAAAATATGATGGGGGATTGGTTGAATATATCCACCACCTAAACAACGAAAAAGAACCACTTTTTAATGAAGTTATTGCTGATGAAAAAACTGAAACTGTAAAAGCTGTTAATCGTGATGAAAACTACACAGTAAAGGTTGAAGTTGCTTTTCAATATAACAAAACATACAACCAATCAATTTTCAGTTTTTGTAACAACATTAATACTACAGAAGGTGGAACCCATGTGGAAGGTTTTCGTAATGCACTTGTTAAGATCATTAATCGCTTTGCTGTTGAAAATAAATTCCTAAAAGATAGTGATGAAAAGATTAACCGTGATGATGTTTGTGAAGGATTAACTGCTATTATTTCCATTAAACACCCAAACCCACAATATGAAGGACAAACTAAAAAGAAGTTAGGTAATACTGAGGTAAGACCTTTAGTTAATAGTGTTGTTAGTGAAATCTTTGAACGCTTCATGTTAGAAAACCCACAAGAAGCAAACGCTATCATCAGAAAAACACTTTTAGCTCAAGAAGCGAGAAGAAGAAGTCAAGAGGCTAGGGAGTTAACTCGTCGTAAATCACCTTTTGATAGTGGTTCATTACCAGGTAAATTAGCTGATTGTACAACCAGAGATCCTTCGATTAGTGAACTTTACATTGTTGAGGGTGATAGTGCTGGTGGCACTGCTAAAACAGGAAGAGATCGTTATTTTCAAGCTATCTTACCCTTAAGAGGAAAGATTTTAAACGTTGAAAAATCTAACTTTGAACAAATCTTTAATAATGCAGAAATTTCTGCATTAGTGATGGCAATAGGCTGTGGGATTAAACCTGATTTTGAACTTGAAAAACTTAGATATAGCAAGATTGTGATCATGACAGATGCTGATGTTGATGGTGCACACATAAGAACACTTCTCTTAACTTTCTTTTTTCGCTTTATGTATCCTTTGGTTGAACAAGGCAATATTTTTATTGCTCAACCCCCACTTTATAAAGTGTCATATTCCCATAAGGATTTATACATGCACACTGATGTTCAACTTGAACAGTGAAAAAGTCAAAACCCTAACGTAAAGTTTGGGTTACAAAGATATAAAGGACTTGGAGAAATGGATGCATTGCAGCTGTGAGAAACAACAATGGATCCTAAGGTTAGAACATTGTTAAAAGTTACTGTTGAAGATGCTTCTATTGCTGATAAAGCTTTTTCACTGTTGATGGGTGATGAAGTTCCCCCAAGAAGAGAATTTATTGAAAAAAATGCTCGTAGTGTTAAAAACATTGATATTTAA 

>Myco_7294_8547_+ 
ATGTTGGATCCAAACAAATTACGCAATAACTATGATTTCTTTAAAAAGAAACTGTTAGAAAGAAATGTAAATGAGCAATTATTAAATCAGTTTATTCAAACTGATAAACTAATGCGCAAAAACTTGCAACAACTTGAACTTGCTAACCAAAAACAAAGCTTGTTGGCAAAACAAGTTGCTAAGCAAAAAGATAATAAAAAGCTATTAGCTGAATCAAAAGAACTTAAGCAGAAGATTGAAAACTTAAATAATGCTTATAAAGATTCACAAAACATTAGTCAAGATTTACTTCTAAATTTTCCTAATATTGCTCATGAATCAGTTCCTGTTGGTAAAAATGAATCAGCAAACTTAGAACTTCTTAAAGAAGGGAGAAAACCAGTTTTTGATTTCAAACCTTTACCACATCGAGAGTTATGTGAAAAGTTAAATTTAGTTGCTTTTGATAAAGCTACTAAGATTAGTGGAACTAGGTTTGTTGCATATACAGATAAAGCAGCTAAACTACTTAGAGCGATAACTAATCTAATGATTGACCTTAATAAAAGCAAGTATCAAGAATGAAACCTGCCAGTTGTTATTAATGAATTAAGTTTAAGATCAACCGGACAACTACCTAAGTTTAAAGATGATGTTTTTAAACTAGAAAACACCCGTTATTATCTTTCTCCAACTTTAGAGGTACAACTTATCAATTTACATGCTAATGAAATTTTTAATGAAGAAGATTTACCTAAATACTACACTGCAACAGGTATTAACTTTCGTCAAGAAGCGGGTAGTGCTGGTAAACAAACCAAAGGAACTATTAGATTGCATCAGTTTCAAAAAACTGAGTTAGTTAAGTTTTGTAAACCTGAAAATGCTATCAATGAATTGGAAGCAATGGTTAGAGATGCTGAACAAATCTTAAAGGCACTTAAGTTACCTTTTAGAAGGTTATTGTTATGTACTGGTGATATGGGCTTTAGTGCTGAAAAAACATATGATCTTGAAGTTTGAATGGCAGCTAGCAATGAATATCGTGAAGTTTCTTCTTGTTCATCTTGTGGTGATTTTCAAGCAAGAAGAGCTATGATTCGTTACAAAGATATTAACAACGGTAAAAACAGTTATGTTGCTACTTTAAATGGAACAGCATTATCTATTGATAGAATTTTTGCTGCAATTCTAGAAAATTTTCAAACAAAAGATGGCAAAATTCTTATCCCACAAGCATTAAAAAAATACCTTGATTTTGACACAATCAAGTAA 
......
 
ORFs_Without_Corresponding_Gene_In_Reference_Metrics:
ATG_Start ,GTG_Start ,TTG_Start ,ATT_Start ,CTG_Start ,Alternative_Start_Codon ,TGA_Stop ,TAA_Stop ,TAG_Stop ,Alternative_Stop_Codon ,Median_Length ,ORFs_on_Positive_Strand ,ORFs_on_Negative_Strand
58.39,17.14,24.47,0.00,0.00,0.00,71.55,20.62,7.83,0.00,287.00,449,356
ORF_Without_Corresponding_Gene_in_Reference:
>Prodigal_1828_2073_+ 
ATGAATCTTTACGATCTTTTAGAACTACCAACTACAGCATCAATAAAAGAAATAAAAATTGCTTATAAAAGATTAGCAAAGCGTTATCACCCTGATGTAAATAAATTAGGTTCGCAAACTTTTGTTGAAATTAATAATGCTTATTCAATATTAAGTGATCCTAACCAAAAGGAAAAATATGATTCAATGCTGAAAGTTAATGATTTTCAAAATCGCATCAAAAATTTAGATATTAGTGTTAGATGA
>Prodigal_2605_2760_+ 
ATGAAAGTAGTTAATAAAGTAAACAAAAGACTGCGTATTTTTTCAAGCTTTTTTGAGAACGATAAATCTAAATTATGGTTCCTTGTTCCAAACGATAAACAAAGTAATCCTAATAAGGGCGTTTTTAACTATAAAACTCAGCACTTTATTGATTAA
>Prodigal_2845_2979_+ 
ATGGAAGAAAATAACAAAGCAAATATCTATGACTCTAGTAGCATTAAGGTCCTTGAAGGACTTGAGGCTGTTAGAAAACGCCCTGGAATGTACATTGGTTCTACTGGCGAAGAAGGTTTGCATCACATGATCTGA
>Prodigal_3010_3255_+ 
ATGGGAGGTTTTGCCAGTTTTGTTAAGCTTACCCTTGAAGATAATTTTGTTACCCGTGTAGAGGATGATGGAAGAGGGATACCTGTTGATATCCATCCTAAGACTAATCGTTCTACAGTTGAAACAGTTTTTACAGTTCTACACGCTGGCGGTAAATTTGATAACGATAGCTATAAAGTGTCAGGTGGTTTACACGGTGTTGGTGCATCAGTTGTTAATGCGCTTAGTTCTTCTTTTAAAGTTTGA
>Prodigal_3319_3513_+ 
TTGGTCCAAGAAGGTAACTCTGAAAAAGAGCATGGAACAATTGTTGAGTTTGTTCCTGATTTCTCTGTAATGGAAAAGAGTGATTACAAACAAACTGTAATTGTAAGCAGACTCCAGCAATTAGCTTTTTTAAACAAGGGAATAAGAATTGACTTTGTTGATAATCGTAAACAAAACCCACAGTCTTTTTCTTGA
>Prodigal_3529_4557_+ 
TTGGTTGAATATATCCACCACCTAAACAACGAAAAAGAACCACTTTTTAATGAAGTTATTGCTGATGAAAAAACTGAAACTGTAAAAGCTGTTAATCGTGATGAAAACTACACAGTAAAGGTTGAAGTTGCTTTTCAATATAACAAAACATACAACCAATCAATTTTCAGTTTTTGTAACAACATTAATACTACAGAAGGTGGAACCCATGTGGAAGGTTTTCGTAATGCACTTGTTAAGATCATTAATCGCTTTGCTGTTGAAAATAAATTCCTAAAAGATAGTGATGAAAAGATTAACCGTGATGATGTTTGTGAAGGATTAACTGCTATTATTTCCATTAAACACCCAAACCCACAATATGAAGGACAAACTAAAAAGAAGTTAGGTAATACTGAGGTAAGACCTTTAGTTAATAGTGTTGTTAGTGAAATCTTTGAACGCTTCATGTTAGAAAACCCACAAGAAGCAAACGCTATCATCAGAAAAACACTTTTAGCTCAAGAAGCGAGAAGAAGAAGTCAAGAGGCTAGGGAGTTAACTCGTCGTAAATCACCTTTTGATAGTGGTTCATTACCAGGTAAATTAGCTGATTGTACAACCAGAGATCCTTCGATTAGTGAACTTTACATTGTTGAGGGTGATAGTGCTGGTGGCACTGCTAAAACAGGAAGAGATCGTTATTTTCAAGCTATCTTACCCTTAAGAGGAAAGATTTTAAACGTTGAAAAATCTAACTTTGAACAAATCTTTAATAATGCAGAAATTTCTGCATTAGTGATGGCAATAGGCTGTGGGATTAAACCTGATTTTGAACTTGAAAAACTTAGATATAGCAAGATTGTGATCATGACAGATGCTGATGTTGATGGTGCACACATAAGAACACTTCTCTTAACTTTCTTTTTTCGCTTTATGTATCCTTTGGTTGAACAAGGCAATATTTTTATTGCTCAACCCCCACTTTATAAAGTGTCATATTCCCATAAGGATTTATACATGCACACTGATGTTCAACTTGAACAGTGA
....
ORFs_Which_Detected_more_than_one_Gene:

GFF 工具：

GFF-加法器：

GFF-Adder 允许将预测的 CDS 添加到现有的参考注释（GFF 或其他工具）中，这会产生包含原始基因的新 GFF 以及来自另一个预测的新 CDS。默认过滤将删除与现有基因重叠超过 50 nt 的额外 CDS。该-gi选项可用于考虑不同的基因组元素，而不仅仅是参考注释中的 CDS。

求助：GFF-Adder -h

usage: GFF-Adder [-h] -dna GENOME_DNA [-rt REFERENCE_TOOL] -ref
                    REFERENCE_ANNOTATION [-gi GENE_IDENT] -at ADDITIONAL_TOOL
                    [-gene_ident GENE_IDENT] -add ADDITIONAL_ANNOTATION
                    [-olap OVERLAP] -o OUTPUT_FILE

optional arguments:
  -h, --help            show this help message and exit
  -dna GENOME_DNA, --genome_DNA GENOME_DNA
                        Genome DNA file (.fa) which both annotations are based
                        on
  -rt REFERENCE_TOOL, --reference_tool REFERENCE_TOOL
                        Which tool format to use as reference? - If not
                        provided, will default to standard Ensembl GFF format,
                        can be Prodigal or any of the other tools available
  -ref REFERENCE_ANNOTATION, --reference_annotation REFERENCE_ANNOTATION
                        Which reference annotation file to use as reference?
  -gi GENE_IDENT, --gene_ident GENE_IDENT
                        Identifier used for extraction of "genic" regions from
                        reference annotation "CDS,rRNA,tRNA": Default for is
                        "CDS"
  -at ADDITIONAL_TOOL, --additional_tool ADDITIONAL_TOOL
                        Which format to use for additional annotation?
  -gene_ident GENE_IDENT
                        Identifier used for identifying genomic features
                        "CDS,rRNA,tRNA"
  -add ADDITIONAL_ANNOTATION, --additional_annotation ADDITIONAL_ANNOTATION
                        Which annotation file to add to reference annotation?
  -olap OVERLAP, --overlap OVERLAP
                        Maximum overlap between reference and additional genic
                        regions (CDS,rRNA etc) - Default: 50 nt
  -o OUTPUT_FILE, --output_file OUTPUT_FILE
                        Output filename

示例：运行 GFF-Adder 以将 Prodial 所做的附加 CDS 预测与来自 Ensembl 的规范注释相结合。

GFF-Adder -dna ~/Testing/Myco.fa -ref ~/Testing/Myco.gff -at Prodigal -add ~/Testing/Prodigal_Myco.gff -o ~/Testing/Myco_Ensembl_GFF_Adder_Prodigal.gff

示例输出：~/ORForise/Testing/Myco_Ensembl_GFF_Adder_Prodigal.gff

##gff-version	3
#	GFF-Adder
#	Run Date:2021-11-10
##Genome DNA File:./Testing/Myco.fa
##Original File: ./Testing/Myco.gff
##Additional File: ./Testing/Prodigal_Myco.gff
.......
Chromosome	Reference_Annotation	CDS	68522	70225	.	-	.	ID=Original_Annotation
Chromosome	Reference_Annotation	CDS	70530	72572	.	+	.	ID=Original_Annotation
Chromosome	Reference_Annotation	CDS	72523	73434	.	+	.	ID=Original_Annotation
Chromosome	Prodigal	CDS	73445	73648	.	+	.	ID=Additional_Annotation
Chromosome	Reference_Annotation	CDS	73690	77685	.	+	.	ID=Original_Annotation
Chromosome	Reference_Annotation	CDS	77685	79085	.	+	.	ID=Original_Annotation
Chromosome	Reference_Annotation	CDS	79089	81035	.	+	.	ID=Original_Annotation
Chromosome	Reference_Annotation	CDS	81046	82596	.	+	.	ID=Original_Annotation
Chromosome	Reference_Annotation	CDS	82620	84044	.	+	.	ID=Original_Annotation
Chromosome	Prodigal	CDS	84082	84312	.	+	.	ID=Additional_Annotation
Chromosome	Prodigal	CDS	84532	84744	.	-	.	ID=Additional_Annotation
Chromosome	Prodigal	CDS	84776	85051	.	+	.	ID=Additional_Annotation

GFF-交叉口：

GFF-Intersector 能够聚合不同的基因组注释和 CDS 预测，并创建一个代表两个现有注释的交集的 GFF。GFF-Intersector 还提供了一个选项，允许保留具有用户定义的差异（最小覆盖率和框架内）的基因。该-gi选项可用于考虑不同的基因组元素，而不仅仅是参考注释中的 CDS。

求助：GFF-Intersector -h

用法： GFF -交叉点 [ - h ]  - dna  GENOME_DNA  [ - rt  REFERENCE_TOOL ]  - ref 
                          REFERENCE_ANNOTATION  [ - gi  GENE_IDENT ]  -在
                          ADDITIONAL_TOOL  -添加 ADDITIONAL_ANNOTATION 
                          [ - cov  COVERAGE ]  - o  OUTPUT_FILE

可选 参数：
  -h ，-- help显示此帮助