介绍：https://vdjtools-doc.readthedocs.io/en/master/

1.下载

对于linux服务器而言，可以直接通过https://github.com/mikessh/vdjtools/releases/tag/1.2.1，进行下载。

zip安装包中包含了所需的jar文件，无需编译，下载即用。
安装相关的R包：

java -jar vdjtools.jar Rinstall

2.使用

事例：

# -p for plotting, -f specifies metadata column for coloring,
# -n tells that factor is continuous
$VDJTOOLS CalcSegmentUsage -m metadata.txt -p -f age -n out/2

输入：

克隆丰度表，对于10x免疫组库的结果而言，可以使用获得的filtered_contig_annotations.csv
对于VDJtools而言，所需的clonotype数据包括这些：

• Variable (V) segment name. V区基因名
• Diversity (D) segment name for some of the receptor chains (TRB, TRD and IGH). Set to . if not aplicable or D segment was not identified. D区基因名，如果是.则是未获得
• Joining (J) segment name. J区基因名
• Complementarity determining region 3 nucleotide sequence (CDR3nt). CDR3 starts with Variable region reference point (conserved Cys residue) and ends with Joining segment reference point (conserved PheTrp). CDR3的核苷酸序列
• Translated CDR3 sequence (CDR3aa). CDR3的氨基酸序列
• Somatic hypermutations (SHMs) in the variable segment (antibody only, planned). V区的体细胞突变

其他都是可选项目。
输入数据应以以制表符分隔的表格形式提供。

格式转换：

将10x的结果数据转换成需要的格式
10x数据：

将样本处理成需要的格式：

Metadata数据格式如下：

分析模块

基础模块：

汇总统计

• CalcBasicStats Computes summary statistics for samples: read counts, mean clonotype sizes, number of non-functional clonotypes, etc
• CalcSegmentUsage Computes Variable (V) and Joining (J) segment usage profiles
• CalcSpectratype Computes spectratype, the distribution of clonotype abundance by CDR3 sequence length
• PlotFancySpectratype Plots spectratype explicitly showing top N clonotypes
• PlotFancyVJUsage Plots the frequency of different V-J pairings
• PlotSpectratypeV Plots distribution of V segment abundance by resulting CDR3 sequence length

多样性评估

丰富和多样性

• PlotQuantileStats Visualizes repertoire clonality
• RarefactionPlot Performs rarefaction analysis
• CalcDiversityStats Computes repertoire diversity estimates

样本间overlap

样本间共享的克隆型

• OverlapPair Computes intersection between a pair of samples
• CalcPairwiseDistances Computes pairwise intersections for a list of samples
• ClusterSamples Performs sample clusterization according to the results of batch intersection
• TrackClonotypes Time-course analysis for a sequence of samples

Pre-processing

过滤和重采样

• Correct Performs a frequency-based erroneous clonotype correction
• Decontaminate Filters possible cross-sample contaminations in a set of samples
• DownSample Performs down-sampling, i.e. takes a subset of random reads from sample(s)
• FilterNonFunctional Filters non-functional clonotypes
• SelectTop Selects a fixed number of top (most abundant) clonotypes from sample(s)
• FilterByFrequency Filters clonotypes based on a specified frequency threshold.
• ApplySampleAsFilter Filters clonotypes that are present in a specified sample from sample(s)
• FilterBySegment Filters clonotypes according to their V/D/J segment

Clonotype表格操作

Clonotype table operations

• PoolSamples Pools clonotypes from several samples together
• JoinSamples Joins a set of samples and generates clonotype abundance profiles

注释

Functional annotation of clonotype tables (antigen specificity, amino acid properties, etc)

• CalcCdrAAProfile Builds a profile of CDR3 regions (V germline, V-D junction, …) using a set of amino-acid physical properties
• Annotate Computes a set of basic (insert size, …) and amino acid physical properties (GRAVY, …) for clonotypes

使用—plot-type png参数获得png格式