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In [1]:



    


source("http://bioconductor.org/biocLite.R")
biocLite("rhdf5")



    


















    






Installing package into ‘/home/ilya/R/x86_64-pc-linux-gnu-library/3.3’
(as ‘lib’ is unspecified)
Bioconductor version 3.3 (BiocInstaller 1.22.3), ?biocLite for help
BioC_mirror: https://bioconductor.org
Using Bioconductor 3.3 (BiocInstaller 1.22.3), R 3.3.1 (2016-06-21).
Installing package(s) ‘rhdf5’
also installing the dependency ‘zlibbioc’

Old packages: 'boot', 'MASS', 'spatial'

















In [1]:



    


devtools::install_github("pachterlab/sleuth")



    


















    






Downloading GitHub repo pachterlab/sleuth@master
from URL https://api.github.com/repos/pachterlab/sleuth/zipball/master
Installing sleuth
Installing shiny
Installing httpuv
'/usr/lib/R/bin/R' --no-site-file --no-environ --no-save --no-restore --quiet  \
  CMD INSTALL '/tmp/RtmpMWwW2H/devtools4d42622c25bc/httpuv'  \
  --library='/home/ilya/R/x86_64-pc-linux-gnu-library/3.3' --install-tests 

Installing R6
'/usr/lib/R/bin/R' --no-site-file --no-environ --no-save --no-restore --quiet  \
  CMD INSTALL '/tmp/RtmpMWwW2H/devtools4d4245d9d26e/R6'  \
  --library='/home/ilya/R/x86_64-pc-linux-gnu-library/3.3' --install-tests 

'/usr/lib/R/bin/R' --no-site-file --no-environ --no-save --no-restore --quiet  \
  CMD INSTALL '/tmp/RtmpMWwW2H/devtools4d428a68d88/shiny'  \
  --library='/home/ilya/R/x86_64-pc-linux-gnu-library/3.3' --install-tests 

'/usr/lib/R/bin/R' --no-site-file --no-environ --no-save --no-restore --quiet  \
  CMD INSTALL  \
  '/tmp/RtmpMWwW2H/devtools4d421e6f6ad4/pachterlab-sleuth-048f055'  \
  --library='/home/ilya/R/x86_64-pc-linux-gnu-library/3.3' --install-tests 


















In [2]:



    


library("sleuth")



    


















    






Loading required package: ggplot2
Loading required package: dplyr

Attaching package: ‘dplyr’

The following objects are masked from ‘package:stats’:

    filter, lag

The following objects are masked from ‘package:base’:

    intersect, setdiff, setequal, union


















In [3]:



    


base_dir <- "../data/kallisto"
sample_id <- dir(file.path(base_dir), pattern='^lo')
sample_id



    


















    







    
	
  1. 'lo03'
    2. 
	
  2. 'lo05'
    3. 
	
  3. 'lo09'
    4. 
	
  4. 'lo11'
    5. 



















In [4]:



    


kal_dirs <- sapply(sample_id, function(id) file.path(base_dir, id))
kal_dirs



    


















    








	
lo03

		
'../data/kallisto/lo03'

	
lo05

		
'../data/kallisto/lo05'

	
lo09

		
'../data/kallisto/lo09'

	
lo11

		
'../data/kallisto/lo11'




















In [5]:



    


dir(kal_dirs)



    


















    







    
	
  1. 'abundance.h5'
    2. 
	
  2. 'abundance.h5'
    3. 
	
  3. 'abundance.h5'
    4. 
	
  4. 'abundance.h5'
    5. 
	
  5. 'abundance.tsv'
    6. 
	
  6. 'abundance.tsv'
    7. 
	
  7. 'abundance.tsv'
    8. 
	
  8. 'abundance.tsv'
    9. 
	
  9. 'run_info.json'
    10. 
	
  10. 'run_info.json'
    11. 
	
  11. 'run_info.json'
    12. 
	
  12. 'run_info.json'
    13. 



















In [7]:



    


gsy_samples <- c('lo05', 'lo09', 'lo03', 'lo11')
gsy_samples



    


















    







    
	
  1. 'lo05'
    2. 
	
  2. 'lo09'
    3. 
	
  3. 'lo03'
    4. 
	
  4. 'lo11'
    5. 



















In [16]:



    


gsy_dirs <- c(kal_dirs[seq(1, length(kal_dirs), 2)], kal_dirs[seq(2, length(kal_dirs), 2)])
gsy_dirs



    


















    








	
lo03

		
'../data/kallisto/lo03'

	
lo09

		
'../data/kallisto/lo09'

	
lo05

		
'../data/kallisto/lo05'

	
lo11

		
'../data/kallisto/lo11'




















In [ ]:



    


gsy



    











In [17]:



    


s2c <- data.frame(sample=c('lo03', 'lo05', 'lo09', 'lo11'), condition=c('gsy', 'wt', 'wt', 'gsy'), path=c('../data/kallisto/lo03','../data/kallisto/lo05', '../data/kallisto/lo09','../data/kallisto/lo11'))
s2c



    


















    









sampleconditionpath


	
lo03                 gsy                  ../data/kallisto/lo03

	
lo05                 wt                   ../data/kallisto/lo05

	
lo09                 wt                   ../data/kallisto/lo09

	
lo11                 gsy                  ../data/kallisto/lo11





















In [ ]:



    






    











In [18]:



    


so <- sleuth_prep(s2c, ~ condition)



    


















    






reading in kallisto results
.









    






Error: is(path, "character") is not TRUE
Traceback:

1. sleuth_prep(s2c, ~condition)
2. lapply(seq_along(kal_dirs), function(i) {
 .     nsamp <- dot(nsamp)
 .     path <- kal_dirs[i]
 .     suppressMessages({
 .         kal <- read_kallisto(path, read_bootstrap = TRUE, max_bootstrap = max_bootstrap)
 .     })
 .     kal$abundance <- dplyr::mutate(kal$abundance, sample = sample_to_covariates$sample[i])
 .     kal
 . })
3. FUN(X[[i]], ...)
4. suppressMessages({
 .     kal <- read_kallisto(path, read_bootstrap = TRUE, max_bootstrap = max_bootstrap)
 . })
5. withCallingHandlers(expr, message = function(c) invokeRestart("muffleMessage"))
6. read_kallisto(path, read_bootstrap = TRUE, max_bootstrap = max_bootstrap)
7. stopifnot(is(path, "character"))
8. stop(sprintf(ngettext(length(r), "%s is not TRUE", "%s are not all TRUE"), 
 .     ch), call. = FALSE, domain = NA)
















In [36]:



    


so <- sleuth_fit(so)



    


















    






fitting measurement error models
shrinkage estimation
Adding missing grouping variables: `x_group`
computing variance of betas

















In [37]:



    


so <- sleuth_fit(so, ~1, 'reduced')



    


















    






fitting measurement error models
shrinkage estimation
Adding missing grouping variables: `x_group`
computing variance of betas

















In [38]:



    


#so <- sleuth_test(so, which_beta='conditionwt')
so <- sleuth_lrt(so, 'reduced', 'full')



    











In [39]:



    


models(so)



    


















    






[  full  ]
formula:  ~condition 
coefficients:
	(Intercept)
 	conditionwt
[  reduced  ]
formula:  ~1 
coefficients:
	(Intercept)

















In [41]:



    


source("http://bioconductor.org/biocLite.R")
biocLite("biomaRt")



    


















    






Bioconductor version 3.3 (BiocInstaller 1.22.3), ?biocLite for help
BioC_mirror: https://bioconductor.org
Using Bioconductor 3.3 (BiocInstaller 1.22.3), R 3.3.1 (2016-06-21).
Installing package(s) ‘biomaRt’
also installing the dependencies ‘BiocGenerics’, ‘Biobase’, ‘IRanges’, ‘RSQLite’, ‘S4Vectors’, ‘XML’, ‘AnnotationDbi’

Old packages: 'boot', 'MASS', 'spatial'

















In [42]:



    


mart <- biomaRt::useMart(biomart = "ENSEMBL_MART_ENSEMBL",
  dataset = "celegans_gene_ensembl",
  host = 'ensembl.org')



    











In [43]:



    


t2g <- biomaRt::getBM(attributes = c("ensembl_transcript_id", "ensembl_gene_id",
    "external_gene_name"), mart = mart)
t2g <- dplyr::rename(t2g, target_id = ensembl_transcript_id,
  ens_gene = ensembl_gene_id, ext_gene = external_gene_name)
so <- sleuth_prep(s2c, ~ condition, target_mapping = t2g)



    


















    






reading in kallisto results
......
normalizing est_counts
19296 targets passed the filter
normalizing tpm
merging in metadata
normalizing bootstrap samples
summarizing bootstraps

















In [44]:



    


so <- sleuth_fit(so)



    


















    






fitting measurement error models
shrinkage estimation
Adding missing grouping variables: `x_group`
computing variance of betas

















In [46]:



    


so <- sleuth_fit(so, ~1, 'reduced')



    


















    






fitting measurement error models
shrinkage estimation
Adding missing grouping variables: `x_group`
computing variance of betas

















In [47]:



    


so <- sleuth_lrt(so, 'reduced', 'full')



    











In [51]:



    


so <- sleuth_wt(so, which_beta = 'conditionwt')



    











In [52]:



    


sleuth_live_settings(test_type='wt')



    


















    






$test_type = 'wt'
















In [ ]:



    


sleuth_live(so)



    


















    






Listening on http://127.0.0.1:42427
Warning message:
: Removed 8 rows containing missing values (geom_point).Warning message:
: Removed 8 rows containing missing values (geom_point).
















In [10]:



    


write.table(res,file='../kallisto/results.tsv', sep='\t')



    











In [ ]:



    


sleuth_live(so)



    


















    






Loading required package: shiny

Listening on http://127.0.0.1:7861

















In [ ]:



    


res <- sleuth_results(so, 'conditionwt')



    











In [ ]:

Content source: eco32i/biodata

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