├── figures ├── quals_beeswarm_9.4_none_100Kbp.png ├── quals_beeswarm_9.4_none_250Kbp.png ├── quals_beeswarm_9.4_none_500Kbp.png ├── quals_beeswarm_10.3_medaka_100Kbp.png ├── quals_beeswarm_10.3_medaka_250Kbp.png ├── quals_beeswarm_10.3_medaka_500Kbp.png ├── quals_beeswarm_10.3_none_100Kbp.png ├── quals_beeswarm_10.3_none_250Kbp.png ├── quals_beeswarm_10.3_none_500Kbp.png ├── quals_beeswarm_9.4_medaka_100Kbp.png ├── quals_beeswarm_9.4_medaka_250Kbp.png └── quals_beeswarm_9.4_medaka_500Kbp.png ├── Snakefile ├── .gitignore ├── scripts └── q_window.R ├── README.md └── LICENSE /figures/quals_beeswarm_9.4_none_100Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_9.4_none_100Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_9.4_none_250Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_9.4_none_250Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_9.4_none_500Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_9.4_none_500Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_10.3_medaka_100Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_10.3_medaka_100Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_10.3_medaka_250Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_10.3_medaka_250Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_10.3_medaka_500Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_10.3_medaka_500Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_10.3_none_100Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_10.3_none_100Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_10.3_none_250Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_10.3_none_250Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_10.3_none_500Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_10.3_none_500Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_9.4_medaka_100Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_9.4_medaka_100Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_9.4_medaka_250Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_9.4_medaka_250Kbp.png -------------------------------------------------------------------------------- /figures/quals_beeswarm_9.4_medaka_500Kbp.png: -------------------------------------------------------------------------------- https://raw.githubusercontent.com/osilander/bonito_benchmarks/HEAD/figures/quals_beeswarm_9.4_medaka_500Kbp.png -------------------------------------------------------------------------------- /Snakefile: -------------------------------------------------------------------------------- 1 | #wildcard_constraints: 2 | # sample=".+10.+" 3 | 4 | # find all fasta files that match fasta from a specific date 5 | STRAINS, = glob_wildcards("data/{sample,.+10.+}.fastq") 6 | # assemble using a couple methods; first flye and raven 7 | ASSEMBLY = ["flye", "raven"] 8 | POLISH = ['medaka', 'none'] 9 | 10 | rule all: 11 | input: 12 | expand("results/{sample}-{assembly}-{polish}.snps", sample=STRAINS, assembly=ASSEMBLY, polish=POLISH) 13 | 14 | rule assemble: 15 | input: 16 | "data/{sample}.fastq" 17 | output: 18 | "results/{sample}/{assembly}/assembly.fasta" 19 | params: 20 | dir="results/{sample}/{assembly}", 21 | size="4.5m" 22 | run: 23 | if wildcards.assembly == 'flye': 24 | shell("flye --nano-raw {input} --out-dir {params.dir} --genome-size {params.size} --threads 24") 25 | if wildcards.assembly == 'raven': 26 | shell("raven {input} -t 12 > {output}") 27 | 28 | rule polish: 29 | input: 30 | nanopore="data/{sample}.fastq", 31 | assembly="results/{sample}/{assembly}/assembly.fasta" 32 | output: 33 | "results/{sample}/{assembly}-{polish}/consensus.fasta" 34 | params: 35 | #model="r941_min_high_g360", 36 | model="r103_min_high_g360", 37 | outdir="results/{sample}/{assembly}-{polish}" 38 | run: 39 | if wildcards.polish == 'medaka': 40 | shell("medaka_consensus -m {params.model} -d {input.assembly} -i {input.nanopore} -o {params.outdir}") 41 | if wildcards.polish == 'none': 42 | # just rename 43 | shell("cp {input.assembly} {output}") 44 | 45 | rule dnadiff: 46 | input: 47 | assembly="results/{sample}/{assembly}-{polish}/consensus.fasta" 48 | output: 49 | "results/{sample}-{assembly}-{polish}.snps" 50 | params: 51 | prefix="results/{sample}-{assembly}-{polish}", 52 | #reference="data/refs/MG1655_U00096.fa" 53 | reference="data/refs/fixed_lab_MG1655_final2.fa" 54 | shell: 55 | """ 56 | dnadiff {params.reference} {input} -p {params.prefix} 57 | """ 58 | 59 | rule plot_quals: 60 | input: 61 | "results/{sample}" 62 | output: 63 | "results/{strain}/beeswarm_quals.pdf" 64 | shell: 65 | "R --slave --no-restore --file=scripts/beeswarm_qual.R --args {input} {output}" 66 | -------------------------------------------------------------------------------- /.gitignore: -------------------------------------------------------------------------------- 1 | # local files 2 | data/ 3 | results/ 4 | .snakemake/ 5 | .snakemake/locks 6 | *R.swp 7 | *.swp 8 | # added by raven 9 | raven.cereal 10 | 11 | # Byte-compiled / optimized / DLL files 12 | __pycache__/ 13 | *.py[cod] 14 | *$py.class 15 | 16 | # C extensions 17 | *.so 18 | 19 | # Distribution / packaging 20 | .Python 21 | build/ 22 | develop-eggs/ 23 | dist/ 24 | downloads/ 25 | eggs/ 26 | .eggs/ 27 | lib/ 28 | lib64/ 29 | parts/ 30 | sdist/ 31 | var/ 32 | wheels/ 33 | pip-wheel-metadata/ 34 | share/python-wheels/ 35 | *.egg-info/ 36 | .installed.cfg 37 | *.egg 38 | MANIFEST 39 | 40 | # PyInstaller 41 | # Usually these files are written by a python script from a template 42 | # before PyInstaller builds the exe, so as to inject date/other infos into it. 43 | *.manifest 44 | *.spec 45 | 46 | # Installer logs 47 | pip-log.txt 48 | pip-delete-this-directory.txt 49 | 50 | # Unit test / coverage reports 51 | htmlcov/ 52 | .tox/ 53 | .nox/ 54 | .coverage 55 | .coverage.* 56 | .cache 57 | nosetests.xml 58 | coverage.xml 59 | *.cover 60 | *.py,cover 61 | .hypothesis/ 62 | .pytest_cache/ 63 | 64 | # Translations 65 | *.mo 66 | *.pot 67 | 68 | # Django stuff: 69 | *.log 70 | local_settings.py 71 | db.sqlite3 72 | db.sqlite3-journal 73 | 74 | # Flask stuff: 75 | instance/ 76 | .webassets-cache 77 | 78 | # Scrapy stuff: 79 | .scrapy 80 | 81 | # Sphinx documentation 82 | docs/_build/ 83 | 84 | # PyBuilder 85 | target/ 86 | 87 | # Jupyter Notebook 88 | .ipynb_checkpoints 89 | 90 | # IPython 91 | profile_default/ 92 | ipython_config.py 93 | 94 | # pyenv 95 | .python-version 96 | 97 | # pipenv 98 | # According to pypa/pipenv#598, it is recommended to include Pipfile.lock in version control. 99 | # However, in case of collaboration, if having platform-specific dependencies or dependencies 100 | # having no cross-platform support, pipenv may install dependencies that don't work, or not 101 | # install all needed dependencies. 102 | #Pipfile.lock 103 | 104 | # PEP 582; used by e.g. github.com/David-OConnor/pyflow 105 | __pypackages__/ 106 | 107 | # Celery stuff 108 | celerybeat-schedule 109 | celerybeat.pid 110 | 111 | # SageMath parsed files 112 | *.sage.py 113 | 114 | # Environments 115 | .env 116 | .venv 117 | env/ 118 | venv/ 119 | ENV/ 120 | env.bak/ 121 | venv.bak/ 122 | 123 | # Spyder project settings 124 | .spyderproject 125 | .spyproject 126 | 127 | # Rope project settings 128 | .ropeproject 129 | 130 | # mkdocs documentation 131 | /site 132 | 133 | # mypy 134 | .mypy_cache/ 135 | .dmypy.json 136 | dmypy.json 137 | 138 | # Pyre type checker 139 | .pyre/ 140 | -------------------------------------------------------------------------------- /scripts/q_window.R: -------------------------------------------------------------------------------- 1 | # this script desperately needs some cleaning 2 | # Rscript --slave --no-restore q_window.R none 3 | 4 | library(beeswarm) 5 | library(tidyr) 6 | library(here) 7 | 8 | args = commandArgs(trailingOnly=TRUE) 9 | plot.pattern <- paste(args[1],".+",args[2],".snps",sep="") 10 | 11 | setwd(here()) 12 | steps <- c(1e5, 2.5e5, 5e5) 13 | steps.lit <- c("100Kbp", "250Kbp", "500Kbp") 14 | 15 | # a buncch of manipulations to simplify the filenames 16 | files <- dir("./results", pattern=plot.pattern) 17 | names <- gsub(".snps","",files) 18 | names <- gsub("-","\n",names) 19 | names <- gsub("K12_","",names) 20 | names <- gsub("_b0.3","\nb0.3.5",names) 21 | names <- gsub("_g4.5","\ng4.5",names) 22 | # print out just to check 23 | #cat(names,"\n") 24 | for (s in 1:length(steps)) { 25 | 26 | # make sure this limit is close to correct so that 27 | # the last window is full length 28 | wins <- seq(1,4.6e6,by=steps[s]) 29 | 30 | quals <- matrix(nrow=5e6/steps[s]-1,ncol=length(files)) 31 | colnames(quals) <- names 32 | med.quals <- vector() 33 | 34 | # loop over all files 35 | for(f in 1:length(files)) { 36 | input <- files[f] 37 | cat(names[f],"\n") 38 | d <- read.table(file=paste("results/",input,sep="")) 39 | q.win <- vector() 40 | 41 | for (i in 1:(length(wins)-1)) { 42 | snps <- subset(d, d[,1] >= wins[i] & d[,1] < wins[i+1]) 43 | cat(wins[i],"\t",dim(snps)[1],"\n") 44 | # it's not obvious how too caluclate this 45 | # if there are no errors 46 | # in the case of no errors, I just add one to the max score 47 | if(dim(snps)[1] > 0) { 48 | q <- max(1,dim(snps)[1]) 49 | quals[i,f] <- -log10(q/steps[s])*10 50 | } 51 | else { quals[i,f] <- -log10(1/steps[s])*10 + 1 } 52 | } 53 | med.quals[f] <- median(quals[,f]) 54 | } 55 | 56 | # we could order by median but perhaps not 57 | quals.df <- as.data.frame(quals) 58 | 59 | # make the matrix long and use factors (for plotting sake) 60 | quals.long <- gather(quals.df, key="Assembler",value="qscore") 61 | 62 | # make the figures directory if needed 63 | ifelse(!dir.exists(file.path("./", "figures")), dir.create(file.path("./", "figures")), FALSE) 64 | 65 | # open a png 66 | png.name <- paste("figures/quals_beeswarm_",args[1],"_",args[2],"_",steps.lit[s],".png",sep="") 67 | png(file=png.name, width=100*length(files),height=480) 68 | 69 | # change some defaults 70 | par(las=1) 71 | par(mar=c(8,5,2,1)) 72 | 73 | # get the beeswarm plot 74 | beeswarm(qscore ~ Assembler, data = quals.long, 75 | log = FALSE, pch = 16, col = rainbow(8), 76 | main = (paste("qscore -",steps.lit[s], "window", sep=" ")), cex.main=1.2, xlab="basecaller-assembler-polisher") 77 | 78 | # add a few lines 79 | abline(h=seq(30,55,by=5),lty=2,lwd=0.5,col="grey",font.main=1) 80 | dev.off() 81 | } 82 | -------------------------------------------------------------------------------- /README.md: -------------------------------------------------------------------------------- 1 | # Nanopore basecallers and assemblers 2 | ## Benchmarking basecallers and assemblers using *E. coli* K12 3 | 4 | This repo compares *E. coli* K12 assemblies (for now, [raven](https://github.com/lbcb-sci/raven) 5 | and [flye](https://github.com/fenderglass/Flye)) 6 | using data from different flowcells chemistries (9.4 and 10.3) and 7 | basecallers (`guppy 4.5.2` and `bonito 0.3.5` [here](https://github.com/nanoporetech/bonito)). 8 | One set of 9.4 `guppy` 9 | basecalls have been filtered (*hq*) using [filtlong](https://github.com/rrwick/Filtlong) 10 | to retain the highest quality 11 | reads (quality weighting of 30 to retain at least 500Mbp) using 12 | `filtlong --mean_q_weight 30 --target_bases 500000000 myreads.fastq > myreads_hq.fastq`. 13 | For the 10.3 chemistry, we did not have enough data to filter. 14 | Note that `bonito` does not produce quality scores (although there may be 15 | ways around this such that read q-scores can be inferred). 16 | 17 | Each assembly is polished using [medaka](https://github.com/nanoporetech/medaka) (or not). 18 | 19 | The results of each assembly is compared to the K12 reference using `dnadiff` 20 | from [mummer](https://github.com/garviz/MUMmer), 21 | and the quality of the assembly is assessed by 22 | calculating the SNPs (i.e. short indels and subsitutions) per window across the genome. 23 | This is plotted as a phred-based q-score 24 | (e.g. q50 means 1 error every 100,000 base pairs). 25 | 26 | ### Notes on data 27 | 28 | The 9.4 data was produced several years ago by our lab. 29 | 30 | The 10.3 data was kindly pointed out to me by [Mari Miyamoto](https://twitter.com/marimiya_tky) 31 | and was collected 32 | by [Rasmus Kirkegaard](https://github.com/Kirk3gaard) in the 33 | [Albertsen lab](https://albertsenlab.org), with an analysis in 34 | an accompanying blogpost [here](https://albertsenlab.org/we-ar10-3-pretty-close-now/) 35 | 36 | ### Notes on errors 37 | For most plots below, I plot errors in the assembly over 250Kbp windows (or 500Kbp). 38 | In a 250 Kbp window, the maximum q-score is 54 ( log10(2.5e5)\*10 ). (actually this 39 | is the score for a window with one error. Below, for windows with no errors, I assign 40 | a q-score of the (max + 1)). 41 | Thus, within 250Kbp-windows, q-scores and associated errors would be: 42 | 43 | | q-score | errors per 250Kbp | 44 | |:--------|:------------------| 45 | | 55 | 0 | 46 | | 54 | 1 | 47 | | 51 | 2 | 48 | | 47 | 5 | 49 | | 44 | 10 | 50 | | 41 | 20 | 51 | | 37 | 50 | 52 | | 34 | 100 | 53 | | 27 | 500 | 54 | 55 | ## 9.4 Flowcells 56 | 57 | ### No polishing 58 | First, the results for 9.4 flowcells with no polishing at all ("none"). Each point in the plot 59 | below indicates the quality score within a 250 Kbp window. `bonito 0.3.5` is 60 | indicated as *b0.3.5*; `guppy 4.5.2` is indicated as *g4.5*. There are two 61 | models for guppy, *fast* (fast) amd *high accuracy* (hac). In addition, for 62 | each set of `guppy` basecalls with `.fastq` scores, there is a matching dataset 63 | that has been filtered for high quality (hq) reads using `filtlong` as indicated above. 64 | 65 | The `flye` assembly basecalled with `bonito` is 66 | the clear winner. However, during `flye` assembly there is a polishing step built-in, so it's noit quite 67 | a fair competition. 68 | All the assemblies using fast basecalling are relatively poor, with q-scores well below 30. The *hac* reads 69 | are considerably higher, and the *hac* reads filtered for quality by `filtlong` are a tad higher still. 70 | 71 | ![beeswarm_K12](figures/quals_beeswarm_9.4_none_250Kbp.png) 72 | 73 | Given the relatively large number of errors for these unpolished genomes, it's also possible to plot 74 | q-scores over 100Kbp pair windows, shown below. This yields slightly more information on the relative 75 | quality of different assemblies as there are more points for each. 76 | 77 | ![beeswarm_K12](figures/quals_beeswarm_9.4_none_100Kbp.png) 78 | 79 | ### With polishing 80 | With `medaka` polishing, the `flye` and `raven` `bonito`-basecalled assemblies come 81 | out *very very* close, at around q49 (3 errors every 250Kbp). Notably, there is one 250Kbp windows in 82 | the `raven` assembly that has *no errors at all* (q55), and two windows that have one error (q54). 83 | 84 | Also interesting is that the `guppy` basecalled assemblies do not 85 | come far behind the `bonito` assemblies, around 46 (6 errors every 250Kbp), 86 | and perhaps slightly higher for the `filtlong`-filtered data (compare, 87 | especially, the `flye` assemblies polished with `medaka` for the 88 | unfiltered and `filtlong` filtered 89 | *hq* data. However, this 90 | means that on average, for a 5Mbp genome, there are close to 150 91 | errors (with the vast majority being indels). 92 | This contrasts with the `bonito` assemblies, which are closer to 93 | 60 errors - and with two 250Kbp windows containing more than 30 of those errors. 94 | 95 | Again, it is clear that the fast basecalled assemblies end up far below all others, 96 | with q-scores around 34 (100 errors per 250Kbp). 97 | 98 | ![beeswarm_K12](figures/quals_beeswarm_9.4_medaka_250Kbp.png) 99 | 100 | Here, given the small number of errors in some assemblies, it is 101 | not useful to plot 100Kbp windows, as *many many* of these windows 102 | have no errors and a perfect q-score of 50. Instead, the results 103 | using 500 Kbp windows are shown below. Here, max q-score is 57, and 104 | while there are no windows without errors, in the `flye` assembly 105 | there is one window with 2 errors (q54). 106 | 107 | ![beeswarm_K12](figures/quals_beeswarm_9.4_medaka_500Kbp.png) 108 | 109 | ## 10.3 Flowcells 110 | 111 | ### No polishing 112 | 113 | The 10.3 chemistry has been designed with the thought there will be fewer indels. 114 | Oxford Nanopore have clearly done a good job here - even *unpolished* 115 | `bonito`-basecalled assemblies haved q-scores well above 40, and in the case of `flye` 116 | (which implements its own polishing), the q-scores are well above 45. This "unpolished" 117 | assembly is *approximately equivalent* to the **9.4** `flye` assembly *with* `medaka` polishing. 118 | 119 | ![beeswarm_K12](figures/quals_beeswarm_10.3_none_250Kbp.png) 120 | 121 | ### With polishing 122 | 123 | But the assemblies that really begin to shine are the **10.3** 124 | `medaka`-polished ones (both `bonito` *and* `guppy`, and for both `raven` and `flye`). 125 | For `bonito` basecalls, *six* (`raven`) and *four* (`flye`) 250Kbp windows have 126 | no errors at all (q55), and an additional four have only one error (q54). 127 | Having said that, there is one window in each of these assemblies that has a very large 128 | number of errors, in fact more than any one winodw of the `guppy`-based assemblies. 129 | 130 | Notably, for 131 | `guppy` *high accuracy* (hac) basecalls, a very large fraction of 250Kbp windows also have no or one errors. 132 | 133 | But the *fast* `guppy` basecalls still wallow a bit in the mud, with polishing only pushing 134 | their assembly qualities up a smidge. 135 | 136 | ![beeswarm_K12](figures/quals_beeswarm_10.3_medaka_250Kbp.png) 137 | 138 | Because these assemblies seem so darn accurate, we can also make a plot with 500Kbp windows. 139 | Although now there are no error-free windows, the majority have one (q57), two (q54), or three (q52) 140 | errors, especially in the case of `raven`. 141 | 142 | Finally, it looks like the `raven` assembly, basecalled wqith `bonito`, and polished with `medaka` 143 | is the winning method, at least for this data and this organism. 144 | 145 | A last and important note here (if you have read this far) is that I am not positive that the 10.3 146 | data is from a clone that is identical to the MG1655 reference sequence from Genbank 147 | (looking at you *Rasmus*). For the 9.4 148 | data, I am comparing to a ground truth reference (our lab strains, which in fact differs from the 149 | Genbank reference by about 7 SNPs). 150 | 151 | ![beeswarm_K12](figures/quals_beeswarm_10.3_medaka_500Kbp.png) 152 | -------------------------------------------------------------------------------- /LICENSE: -------------------------------------------------------------------------------- 1 | GNU GENERAL PUBLIC LICENSE 2 | Version 2, June 1991 3 | 4 | Copyright (C) 1989, 1991 Free Software Foundation, Inc., 5 | 51 Franklin Street, Fifth Floor, Boston, MA 02110-1301 USA 6 | Everyone is permitted to copy and distribute verbatim copies 7 | of this license document, but changing it is not allowed. 8 | 9 | Preamble 10 | 11 | The licenses for most software are designed to take away your 12 | freedom to share and change it. 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However, as a 160 | special exception, the source code distributed need not include 161 | anything that is normally distributed (in either source or binary 162 | form) with the major components (compiler, kernel, and so on) of the 163 | operating system on which the executable runs, unless that component 164 | itself accompanies the executable. 165 | 166 | If distribution of executable or object code is made by offering 167 | access to copy from a designated place, then offering equivalent 168 | access to copy the source code from the same place counts as 169 | distribution of the source code, even though third parties are not 170 | compelled to copy the source along with the object code. 171 | 172 | 4. You may not copy, modify, sublicense, or distribute the Program 173 | except as expressly provided under this License. Any attempt 174 | otherwise to copy, modify, sublicense or distribute the Program is 175 | void, and will automatically terminate your rights under this License. 176 | However, parties who have received copies, or rights, from you under 177 | this License will not have their licenses terminated so long as such 178 | parties remain in full compliance. 179 | 180 | 5. You are not required to accept this License, since you have not 181 | signed it. However, nothing else grants you permission to modify or 182 | distribute the Program or its derivative works. These actions are 183 | prohibited by law if you do not accept this License. Therefore, by 184 | modifying or distributing the Program (or any work based on the 185 | Program), you indicate your acceptance of this License to do so, and 186 | all its terms and conditions for copying, distributing or modifying 187 | the Program or works based on it. 188 | 189 | 6. Each time you redistribute the Program (or any work based on the 190 | Program), the recipient automatically receives a license from the 191 | original licensor to copy, distribute or modify the Program subject to 192 | these terms and conditions. You may not impose any further 193 | restrictions on the recipients' exercise of the rights granted herein. 194 | You are not responsible for enforcing compliance by third parties to 195 | this License. 196 | 197 | 7. If, as a consequence of a court judgment or allegation of patent 198 | infringement or for any other reason (not limited to patent issues), 199 | conditions are imposed on you (whether by court order, agreement or 200 | otherwise) that contradict the conditions of this License, they do not 201 | excuse you from the conditions of this License. If you cannot 202 | distribute so as to satisfy simultaneously your obligations under this 203 | License and any other pertinent obligations, then as a consequence you 204 | may not distribute the Program at all. For example, if a patent 205 | license would not permit royalty-free redistribution of the Program by 206 | all those who receive copies directly or indirectly through you, then 207 | the only way you could satisfy both it and this License would be to 208 | refrain entirely from distribution of the Program. 209 | 210 | If any portion of this section is held invalid or unenforceable under 211 | any particular circumstance, the balance of the section is intended to 212 | apply and the section as a whole is intended to apply in other 213 | circumstances. 214 | 215 | It is not the purpose of this section to induce you to infringe any 216 | patents or other property right claims or to contest validity of any 217 | such claims; this section has the sole purpose of protecting the 218 | integrity of the free software distribution system, which is 219 | implemented by public license practices. Many people have made 220 | generous contributions to the wide range of software distributed 221 | through that system in reliance on consistent application of that 222 | system; it is up to the author/donor to decide if he or she is willing 223 | to distribute software through any other system and a licensee cannot 224 | impose that choice. 225 | 226 | This section is intended to make thoroughly clear what is believed to 227 | be a consequence of the rest of this License. 228 | 229 | 8. If the distribution and/or use of the Program is restricted in 230 | certain countries either by patents or by copyrighted interfaces, the 231 | original copyright holder who places the Program under this License 232 | may add an explicit geographical distribution limitation excluding 233 | those countries, so that distribution is permitted only in or among 234 | countries not thus excluded. In such case, this License incorporates 235 | the limitation as if written in the body of this License. 236 | 237 | 9. The Free Software Foundation may publish revised and/or new versions 238 | of the General Public License from time to time. Such new versions will 239 | be similar in spirit to the present version, but may differ in detail to 240 | address new problems or concerns. 241 | 242 | Each version is given a distinguishing version number. If the Program 243 | specifies a version number of this License which applies to it and "any 244 | later version", you have the option of following the terms and conditions 245 | either of that version or of any later version published by the Free 246 | Software Foundation. If the Program does not specify a version number of 247 | this License, you may choose any version ever published by the Free Software 248 | Foundation. 249 | 250 | 10. If you wish to incorporate parts of the Program into other free 251 | programs whose distribution conditions are different, write to the author 252 | to ask for permission. For software which is copyrighted by the Free 253 | Software Foundation, write to the Free Software Foundation; we sometimes 254 | make exceptions for this. Our decision will be guided by the two goals 255 | of preserving the free status of all derivatives of our free software and 256 | of promoting the sharing and reuse of software generally. 257 | 258 | NO WARRANTY 259 | 260 | 11. BECAUSE THE PROGRAM IS LICENSED FREE OF CHARGE, THERE IS NO WARRANTY 261 | FOR THE PROGRAM, TO THE EXTENT PERMITTED BY APPLICABLE LAW. EXCEPT WHEN 262 | OTHERWISE STATED IN WRITING THE COPYRIGHT HOLDERS AND/OR OTHER PARTIES 263 | PROVIDE THE PROGRAM "AS IS" WITHOUT WARRANTY OF ANY KIND, EITHER EXPRESSED 264 | OR IMPLIED, INCLUDING, BUT NOT LIMITED TO, THE IMPLIED WARRANTIES OF 265 | MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE. THE ENTIRE RISK AS 266 | TO THE QUALITY AND PERFORMANCE OF THE PROGRAM IS WITH YOU. SHOULD THE 267 | PROGRAM PROVE DEFECTIVE, YOU ASSUME THE COST OF ALL NECESSARY SERVICING, 268 | REPAIR OR CORRECTION. 269 | 270 | 12. IN NO EVENT UNLESS REQUIRED BY APPLICABLE LAW OR AGREED TO IN WRITING 271 | WILL ANY COPYRIGHT HOLDER, OR ANY OTHER PARTY WHO MAY MODIFY AND/OR 272 | REDISTRIBUTE THE PROGRAM AS PERMITTED ABOVE, BE LIABLE TO YOU FOR DAMAGES, 273 | INCLUDING ANY GENERAL, SPECIAL, INCIDENTAL OR CONSEQUENTIAL DAMAGES ARISING 274 | OUT OF THE USE OR INABILITY TO USE THE PROGRAM (INCLUDING BUT NOT LIMITED 275 | TO LOSS OF DATA OR DATA BEING RENDERED INACCURATE OR LOSSES SUSTAINED BY 276 | YOU OR THIRD PARTIES OR A FAILURE OF THE PROGRAM TO OPERATE WITH ANY OTHER 277 | PROGRAMS), EVEN IF SUCH HOLDER OR OTHER PARTY HAS BEEN ADVISED OF THE 278 | POSSIBILITY OF SUCH DAMAGES. 279 | 280 | END OF TERMS AND CONDITIONS 281 | 282 | How to Apply These Terms to Your New Programs 283 | 284 | If you develop a new program, and you want it to be of the greatest 285 | possible use to the public, the best way to achieve this is to make it 286 | free software which everyone can redistribute and change under these terms. 287 | 288 | To do so, attach the following notices to the program. It is safest 289 | to attach them to the start of each source file to most effectively 290 | convey the exclusion of warranty; and each file should have at least 291 | the "copyright" line and a pointer to where the full notice is found. 292 | 293 | 294 | Copyright (C) 295 | 296 | This program is free software; you can redistribute it and/or modify 297 | it under the terms of the GNU General Public License as published by 298 | the Free Software Foundation; either version 2 of the License, or 299 | (at your option) any later version. 300 | 301 | This program is distributed in the hope that it will be useful, 302 | but WITHOUT ANY WARRANTY; without even the implied warranty of 303 | MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the 304 | GNU General Public License for more details. 305 | 306 | You should have received a copy of the GNU General Public License along 307 | with this program; if not, write to the Free Software Foundation, Inc., 308 | 51 Franklin Street, Fifth Floor, Boston, MA 02110-1301 USA. 309 | 310 | Also add information on how to contact you by electronic and paper mail. 311 | 312 | If the program is interactive, make it output a short notice like this 313 | when it starts in an interactive mode: 314 | 315 | Gnomovision version 69, Copyright (C) year name of author 316 | Gnomovision comes with ABSOLUTELY NO WARRANTY; for details type `show w'. 317 | This is free software, and you are welcome to redistribute it 318 | under certain conditions; type `show c' for details. 319 | 320 | The hypothetical commands `show w' and `show c' should show the appropriate 321 | parts of the General Public License. Of course, the commands you use may 322 | be called something other than `show w' and `show c'; they could even be 323 | mouse-clicks or menu items--whatever suits your program. 324 | 325 | You should also get your employer (if you work as a programmer) or your 326 | school, if any, to sign a "copyright disclaimer" for the program, if 327 | necessary. Here is a sample; alter the names: 328 | 329 | Yoyodyne, Inc., hereby disclaims all copyright interest in the program 330 | `Gnomovision' (which makes passes at compilers) written by James Hacker. 331 | 332 | , 1 April 1989 333 | Ty Coon, President of Vice 334 | 335 | This General Public License does not permit incorporating your program into 336 | proprietary programs. If your program is a subroutine library, you may 337 | consider it more useful to permit linking proprietary applications with the 338 | library. If this is what you want to do, use the GNU Lesser General 339 | Public License instead of this License. 340 | --------------------------------------------------------------------------------