plotQualityProfile

emlas · July 30, 2020, 7:17pm

Hi, This time the error appeared:How to proceed
">" expected at beginning of line 1


> taxa <- assignTaxonomy(seqtab.nochim,
+ "C:/Users/salme/Desktop/SYNCdata/NGS 2019/2423 endo/UNZIP/filtered", multithread=TRUE)
Error in .Call2("fasta_index", filexp_list, nrec, skip, seek.first.rec,  : 
  reading FASTA file C:/Users/salme/Desktop/SYNCdata/NGS 2019/2423 endo/UNZIP/filtered/TA-2423-Rhizo2Ser205-1-ENDO_F_filt.fastq.gz: ">" expected at beginning of line 1

nirgrahamuk · July 30, 2020, 7:26pm

a naive reading of that is that the file being read in is improperly structured.

emlas · July 31, 2020, 3:02am

dirPath: C: /Users/salme/Desktop/SYNCdata/NGS 2019/2423 endo/UNZIP/filtered
the filtered folder contains gz files below.
I understand ">" is missing. WHERE should it be used?

nirgrahamuk · July 31, 2020, 6:12am

inside the file? I dont know

emlas · July 31, 2020, 6:22am

the filtered folder contains the fastq sequences after the R filtrations. They are ALL zipped files. I trust it is correct to place the downloaded sliva__nr_132_train_set.fa file which also is gz file. Should I unzip them? in order to have ">"?

nirgrahamuk · July 31, 2020, 6:52am

Im sorry I dont know. I hope you can find someone with experience with these files and packages to help you. I think I helped you as much as a general R user could. Sorry.

emlas · July 31, 2020, 6:58am

A general question: should I update packages (1), download library's (2) every time I use R?

nirgrahamuk · July 31, 2020, 7:04am

I dont.
But you must use library() to load relevant libraries for your work, thats why library calls are commonly found at the top of each script.

emlas · July 31, 2020, 7:10am

loading libraries. every now and then it says that things are masked (below): Do I need to do smth about it?
Version:1.0 StartHTML:0000000107 EndHTML:0000001291 StartFragment:0000000127 EndFragment:0000001273

library(phangorn) Loading required package: ape Attaching package: ‘ape’ The following object is masked from ‘package:Biostrings’: complement

emlas · July 31, 2020, 7:19am

Q2
I paste below the directory with fastq files and silva. Does it look okay to you?
dir(path="C:/Users/salme/Desktop/SYNCdata/NGS 2019/2423 endo/UNZIP/filtered"

)
[1] "silva_nr_v128_train_set.fa.gz"
[2] "TA-2423-Rhizo2Ser205-1-ENDO_F_filt.fastq.gz"
[3] "TA-2423-Rhizo2Ser205-1-ENDO_R_filt.fastq.gz"
[4] "TA-2423-Rhizo2Ser205-2-ENDO_F_filt.fastq.gz"
[5] "TA-2423-Rhizo2Ser205-2-ENDO_R_filt.fastq.gz"
[6] "TA-2423-Rhizo2Ser205-3-ENDO_F_filt.fastq.gz"
[7] "TA-2423-Rhizo2Ser205-3-ENDO_R_filt.fastq.gz"
[8] "TA-2423-Rhizo2Ser211-1-ENDO_F_filt.fastq.gz"
[9] "TA-2423-Rhizo2Ser211-1-ENDO_R_filt.fastq.gz"
[10] "TA-2423-Rhizo2Ser211-2-ENDO_F_filt.fastq.gz"
[11] "TA-2423-Rhizo2Ser211-2-ENDO_R_filt.fastq.gz"
[12] "TA-2423-Rhizo2Ser211-3-ENDO_F_filt.fastq.gz"
[13] "TA-2423-Rhizo2Ser211-3-ENDO_R_filt.fastq.gz"
[14] "TA-2423-Rhizo2Ser229-1-ENDO_F_filt.fastq.gz"
[15] "TA-2423-Rhizo2Ser229-1-ENDO_R_filt.fastq.gz"
[16] "TA-2423-Rhizo2Ser229-2-ENDO_F_filt.fastq.gz"
[17] "TA-2423-Rhizo2Ser229-2-ENDO_R_filt.fastq.gz"
[18] "TA-2423-Rhizo2Ser229-3-ENDO_F_filt.fastq.gz"
[19] "TA-2423-Rhizo2Ser229-3-ENDO_R_filt.fastq.gz"
[20] "TA-2423-Rhizo2Ser42-1-ENDO_F_filt.fastq.gz"
[21] "TA-2423-Rhizo2Ser42-1-ENDO_R_filt.fastq.gz"
[22] "TA-2423-Rhizo2Ser42-2-ENDO_F_filt.fastq.gz"
[23] "TA-2423-Rhizo2Ser42-2-ENDO_R_filt.fastq.gz"
[24] "TA-2423-Rhizo2Ser42-3-ENDO_F_filt.fastq.gz"
[25] "TA-2423-Rhizo2Ser42-3-ENDO_R_filt.fastq.gz"
[26] "TA-2423-Rhizo2Ser72-1-ENDO_F_filt.fastq.gz"
[27] "TA-2423-Rhizo2Ser72-1-ENDO_R_filt.fastq.gz"
[28] "TA-2423-Rhizo2Ser72-2-ENDO_F_filt.fastq.gz"
[29] "TA-2423-Rhizo2Ser72-2-ENDO_R_filt.fastq.gz"
[30] "TA-2423-Rhizo2Ser72-3-ENDO_F_filt.fastq.gz"
[31] "TA-2423-Rhizo2Ser72-3-ENDO_R_filt.fastq.gz"
[32] "TA-2423-Rhizo2Ser97-1-ENDO_F_filt.fastq.gz"
[33] "TA-2423-Rhizo2Ser97-1-ENDO_R_filt.fastq.gz"
[34] "TA-2423-Rhizo2Ser97-2-ENDO_F_filt.fastq.gz"
[35] "TA-2423-Rhizo2Ser97-2-ENDO_R_filt.fastq.gz"
[36] "TA-2423-Rhizo2Ser97-3-ENDO_F_filt.fastq.gz"
[37] "TA-2423-Rhizo2Ser97-3-ENDO_R_filt.fastq.gz"

emlas · July 31, 2020, 9:30am

I see that there are input, filtered denoised, mergerd and, nochim files. Where are the nochim files located? Apparently I should place my silva file in the folder in order the taxonomy assignment to work. Correct?

nirgrahamuk · July 31, 2020, 9:39am

is the method capable of reading both .fa and fastq files ? I ask because there are both types in that list

emlas · July 31, 2020, 9:44am

I just follow DADA2 pipeline tutorial 1.12
They recommend so

nirgrahamuk · July 31, 2020, 9:54am

I just looked at the tutorial and it shows them passing the path of the

"Training/silva_nr_v132_train_set.fa.gz"

only, they do not list additional files in the param call for assigntaxonomy

emlas · July 31, 2020, 11:25am

Yes, but their former description creates fastq files. And they ask to place the silva file in the directory with the fastq files.
An other issue concerns " multithread=TRUE" Should it be " multithread=FALSE" for the Windows throughout the tutorial ?

nirgrahamuk · July 31, 2020, 11:26am

so maybe its sufficient, to have the file placed where things it references are, but not to name them to the function call...

multithread is an option to run on more than 1 core, it doesnt seem supported on windows, if you are on windows, therefore you shouldnt use it, even if you follow a tutorial that uses it. They are likely on unix if they are using it.

emlas · July 31, 2020, 11:57am

I tried this see below.
taxa <- assignTaxonomy(seqtab.nochim,

"~/silva_nr_v132_train_set.fa.gz", multithread=FALSE)
Error: Input/Output
no input files found
dirPath: C:\Users\salme\Documents/silva_nr_v132_train_set.fa.gz
pattern: character(0)

nirgrahamuk · July 31, 2020, 12:03pm

it seems that you dont have the files in this place.

emlas · July 31, 2020, 12:13pm

yes I know it seems. yet there are files
dir(path="C:/Users/salme/Desktop/SYNCdata/NGS 2019/2423 endo/UNZIP/filtered/")
[1] "silva_nr_v128_train_set.fa.gz"
[2] "TA-2423-Rhizo2Ser205-1-ENDO_F_filt.fastq.gz"
[3] "TA-2423-Rhizo2Ser205-1-ENDO_R_filt.fastq.gz"
[4] "TA-2423-Rhizo2Ser205-2-ENDO_F_filt.fastq.gz"
[5] "TA-2423-Rhizo2Ser205-2-ENDO_R_filt.fastq.gz"
[6] "TA-2423-Rhizo2Ser205-3-ENDO_F_filt.fastq.gz"
[7] "TA-2423-Rhizo2Ser205-3-ENDO_R_filt.fastq.gz"
[8] "TA-2423-Rhizo2Ser211-1-ENDO_F_filt.fastq.gz"
[9] "TA-2423-Rhizo2Ser211-1-ENDO_R_filt.fastq.gz"
[10] "TA-2423-Rhizo2Ser211-2-ENDO_F_filt.fastq.gz"
[11] "TA-2423-Rhizo2Ser211-2-ENDO_R_filt.fastq.gz"
[12] "TA-2423-Rhizo2Ser211-3-ENDO_F_filt.fastq.gz"
[13] "TA-2423-Rhizo2Ser211-3-ENDO_R_filt.fastq.gz"
[14] "TA-2423-Rhizo2Ser229-1-ENDO_F_filt.fastq.gz"
[15] "TA-2423-Rhizo2Ser229-1-ENDO_R_filt.fastq.gz"
[16] "TA-2423-Rhizo2Ser229-2-ENDO_F_filt.fastq.gz"
[17] "TA-2423-Rhizo2Ser229-2-ENDO_R_filt.fastq.gz"
[18] "TA-2423-Rhizo2Ser229-3-ENDO_F_filt.fastq.gz"
[19] "TA-2423-Rhizo2Ser229-3-ENDO_R_filt.fastq.gz"
[20] "TA-2423-Rhizo2Ser42-1-ENDO_F_filt.fastq.gz"
[21] "TA-2423-Rhizo2Ser42-1-ENDO_R_filt.fastq.gz"
[22] "TA-2423-Rhizo2Ser42-2-ENDO_F_filt.fastq.gz"
[23] "TA-2423-Rhizo2Ser42-2-ENDO_R_filt.fastq.gz"
[24] "TA-2423-Rhizo2Ser42-3-ENDO_F_filt.fastq.gz"
[25] "TA-2423-Rhizo2Ser42-3-ENDO_R_filt.fastq.gz"
[26] "TA-2423-Rhizo2Ser72-1-ENDO_F_filt.fastq.gz"
[27] "TA-2423-Rhizo2Ser72-1-ENDO_R_filt.fastq.gz"
[28] "TA-2423-Rhizo2Ser72-2-ENDO_F_filt.fastq.gz"
[29] "TA-2423-Rhizo2Ser72-2-ENDO_R_filt.fastq.gz"
[30] "TA-2423-Rhizo2Ser72-3-ENDO_F_filt.fastq.gz"
[31] "TA-2423-Rhizo2Ser72-3-ENDO_R_filt.fastq.gz"
[32] "TA-2423-Rhizo2Ser97-1-ENDO_F_filt.fastq.gz"
[33] "TA-2423-Rhizo2Ser97-1-ENDO_R_filt.fastq.gz"
[34] "TA-2423-Rhizo2Ser97-2-ENDO_F_filt.fastq.gz"
[35] "TA-2423-Rhizo2Ser97-2-ENDO_R_filt.fastq.gz"
[36] "TA-2423-Rhizo2Ser97-3-ENDO_F_filt.fastq.gz"
[37] "TA-2423-Rhizo2Ser97-3-ENDO_R_filt.fastq.gz"

nirgrahamuk · July 31, 2020, 12:14pm

but C:/Users/salme/Desktop/SYNCdata/NGS 2019/2423 endo/UNZIP/filtered/
and C:\Users\salme\Documents/
are different