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1 =======
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2 ezBAMQC
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3 =======
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4
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5 *"ezBAMQC, a tool to check the quality of mapped next generation sequencing files."*
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6
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7 :Codeology Icon:
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8
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9 .. image:: https://raw.githubusercontent.com/mhammell-laboratory/bamqc/master/doc/bamqc-icon.gif
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10 :alt: generated at codeology.braintreepayments.com/mhammell-laboratory/bamqc
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11 :align: right
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12 :target: http://codeology.braintreepayments.com/mhammell-laboratory/bamqc
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13
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14 :Description:
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15
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16 ezBAMQC is a tool to check the quality of either one or many mapped next-generation-sequencing datasets. It conducts comprehensive evaluations of aligned sequencing data from multiple aspects including: clipping profile, mapping quality distribution, mapped read length distribution, genomic/transcriptomic mapping distribution, inner distance distribution (for paired-end reads), ribosomal RNA contamination, transcript 5’ and 3’ end bias, transcription dropout rate, sample correlations, sample reproducibility, sample variations. It outputs a set of tables and plots and one HTML page that contains a summary of the results. Many metrics are designed for RNA-seq data specifically, but ezBAMQC can be applied to any mapped sequencing dataset such as RNA-seq, CLIP-seq, GRO-seq, ChIP-seq, DNA-seq and so on.
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17
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18 :Links:
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19
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20 `Github Page <https://github.com/mhammell-laboratory/bamqc>`_
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21
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22 `Pypi Page <https://pypi.python.org/pypi/ezBAMQC>`_
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23
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24 `MHammell Lab <http://hammelllab.labsites.cshl.edu/software>`_
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25
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26 :Authors:
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27 Ying Jin, David Molik, and Molly Hammell
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28
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29 :Version: 0.6.7
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30
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31 :Contact:
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32 Ying Jin (yjin@cshl.edu)
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33
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34 Installation guide for ezBAMQC for from source installs
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35 =======================================================
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36
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37 When installing ezBAMQC there are several options, but the main point is: since ezBAMQC uses C++ STD 11 you'll need a version of GCC that can support that, this useally means 4.8 or 4.9. beyond that, you'll need Python, R and Corrplot for interfacing with the C code.
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38
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39 :Intallation:
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40 `Source Code <https://github.com/mhammell-laboratory/ezBAMQC/releases>`_
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41
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42 `Pypi <https://pypi.python.org/pypi?:action=display&name=ezBAMQC>`_
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43
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44 :Prerequisites:
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45 * `python2.7 <https://www.python.org/download/releases/2.7/>`_
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46 * `R <https://www.r-project.org/>`_
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47 * `corrplot <https://cran.r-project.org/web/packages/corrplot/>`_
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48 * `GCC 4.8.1 or greater <https://gcc.gnu.org/gcc-4.8/>`_ GCC 4.9.1 or greater is recomended for PyPi install
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49
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50 :Notes:
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51 * While there are multiple methods of installing the prerequistes it may help to look at (if using a yum based linux distro):*
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52 * `Devtoolset-3 <https://access.redhat.com/documentation/en-US/Red_Hat_Developer_Toolset/3/html/User_Guide/sect-Red_Hat_Developer_Toolset-Install.html>`_ for GCC compilers
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53 * `IUS <https://ius.io/>`_ for Python2.7
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54 * `Software Collections <https://www.softwarecollections.org/>`_ for collections of software (like devtoolset 3 or python)
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55 * `rpmfinder <https://www.rpmfind.net/>`_ for searching rpms across mutliple systems
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56
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57 Setup
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58 =====
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59
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60 1) Make sure that the GCC comiler is in your PATH:
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61
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62 ::
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63
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64 export PATH=/path/to/gcc:$PATH
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65
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66 2) Make sure that python2.7 is in your PYTHONPATH:
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67
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68 ::
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69
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70 export PYTHONPATH=/path/to/python2.7/site-packages:$PYTHONPATH
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71
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72 3) There are three methods of installation of ezBAMQC, from source, from setup.py, and from pypi, once prequistes are setup.
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73
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74 From Source
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75 ~~~~~~~~~~~
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76
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77 1) Download source
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78
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79 2) Unpack tarball and go to the directory of the package:
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80
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81 ::
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82
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83 tar xvfz bamqc-0.6.7.tar.gz
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84
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85 cd bamqc-0.6.7
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86
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87 3) Run make:
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88
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89 ::
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90
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91 make
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92
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93 From Setup.py
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94 ~~~~~~~~~~~~~
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95
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96 ::
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97
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98 python2.7 setup.py install
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99
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100 From Pypi
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101 ~~~~~~~~~
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102
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103 ::
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104
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105 pip2.7 install BAMqc
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106
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107 Usage
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108 =====
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109
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110 ::
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111
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112 ezBAMQC [-h] -i alignment_files [alignment_files ...] -r [refgene]
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113 [-f [attrID]] [--rRNA [rRNA]] -o [dir] [--stranded [stranded]]
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114 [-q [mapq]] [-l labels [labels ...]] [-t NUMTHREADS]
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115
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116 optional arguments:
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117
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118 ::
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119
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120 -h, --help show this help message and exit.
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121 -i, --inputFile alignment files. Could be multiple SAM/BAM files separated by space. Required.
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122 -r, --refgene gene annotation file in GTF format. Required
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123 -f the read summation at which feature level in the GTF file. DEFAULT: gene_id.
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124 --rRNA rRNA coordinates in BED format.
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125 -o, --outputDir output directory. Required.
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126 --stranded strandness of the library?
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127 yes : sense stranded
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128 reverse : reverse stranded
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129 no : not stranded
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130 DEFAULT: yes.
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131 -q, --mapq Minimum mapping quality (phred scaled) for an alignment to be called uniquely mapped. DEFAULT:30
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132 -l, --label Labels of input files. DEFAULT:smp1 smp2 ...
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133 -t, --threads Number of threads to use. DEFAULT:1
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134
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135 Example:
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136
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137 ::
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138
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139 ezBAMQC -i test-data/exp_data/treat1.bam test-data/exp_data/treat2.bam test-data/exp_data/treat3.bam -r test-data/exp_data/hg9_refGene.gtf -q 30 --rRNA test-data/exp_data/hg19_rRNA.bed -o exp_output2
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140
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141 Please find the example output from folder test-data.
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142
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143 FAQ
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144 ===
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145 Q: Why use ezBAMQC?
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146
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147 A: ezBAMQC is efficient and easy to use. With one command line, it reports a comprehensive evaluation of the data with a set of plots and tables.The ability to assess multiple samples together with high efficiency make it especially useful in cases where there are a large number of samples from the same condition, genotype, or treatment. ezBAMQC was written in C++ and supports multithreading. A mouse RNA-seq sample with 120M alignments can be done in 8 minutes with 5 threads.
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148
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149 Q: Why the total number of reads reported by ezBAMQC does not match with samtools flagstat?
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150
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151 A: The difference is because of non-uniquely mapped reads or multiply aligned reads (multi-reads). Samtools flagstat counts each multiple aligment as a different reads, but ezBAMQC counts reads accoriding to the read ID, i.e., each individual read will be counted once no matter that it is a uniquely mapped read or multi-read.
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152
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153 Q: What is "Low Quality Reads" ?
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154
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155 A: Reads marked as qc fail accoriding to SAM format or reads with mapping quality lower than the value set by the option -q will be considered as "Low Quality Reads".
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156
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157 Q: How the setting of option -q alter the results?
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158
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159 A: Reads with low quality, i.e., did not pass -q cutoff, are only counted in Total Reads, Mapped Reads, and Mappability by mapping quality plot. The rest of the report does not include low quality reads.
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160
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161 Q: Do multi-reads (non-uniquely mapped reads) have been considered in Read distribution and gene quantification?
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162
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163 A: No. Only uniquely mapped reads were counted.
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164
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165
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166 Acknowledgements
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167 ================
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168
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169 #) Samtools contributors
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170 #) Users' valuable feedback
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171
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172 Copying & Distribution
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173 ======================
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174
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175 ezBAMQC is free software: you can redistribute it and/or modify
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176 it under the terms of the GNU General Public License as published by
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177 the Free Software Foundation, either version 3 of the License, or
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178 (at your option) any later version.
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179
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180 This program is distributed in the hope that it will be useful,
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181 but *WITHOUT ANY WARRANTY*; without even the implied warranty of
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182 *MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE*. See the
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183 GNU General Public License for more details.
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184
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185 You should have received a copy of the GNU General Public License
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186 along with ezBAMQC. If not, see `this website <http://www.gnu.org/licenses/>`_
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