Hi cao,

Thanks for your great work! I met a question that when I mapped the reads to rRNA using STAR (2.5.3b) configuration the same as yours, the speed is less than 2 M/h and it takes several days and still not finish. I tried to change machines but it didn't work. The load of CPUs is full during the process. Is it normal for RIC-seq data with your configuration or I should check rRNA reference or STAR version or machine again ?

Thanks!

Time Speed Read Read Mapped Mapped Mapped Mapped Unmapped Unmapped Unmapped Unmapped
M/hr number length unique length MMrate multi multi+ MM short other
Dec 05 05:02:03 0.2 1049165 128 1.7% 134.7 1.3% 21.6% 0.0% 0.0% 58.7% 18.0%
Dec 05 05:04:00 0.4 2092755 128 1.7% 134.6 1.4% 21.2% 0.0% 0.0% 59.1% 18.0%
Dec 05 05:07:15 0.5 3135946 128 1.6% 134.4 1.4% 21.0% 0.0% 0.0% 59.3% 18.1%
Dec 05 05:09:53 0.7 4178318 128 1.6% 134.4 1.4% 20.8% 0.0% 0.0% 59.5% 18.1%
Dec 05 05:11:31 1.0 6260778 128 1.6% 134.2 1.4% 20.5% 0.0% 0.0% 59.8% 18.2%
Dec 05 05:12:41 1.2 7302778 128 1.6% 134.1 1.5% 20.4% 0.0% 0.0% 59.8% 18.2%
Dec 05 05:13:41 1.5 9378538 128 1.6% 134.2 1.5% 20.2% 0.0% 0.0% 60.0% 18.2%

Hi,
Thanks for your work! I met a question. In your article, you wrote that all the mapped reads from STAR and bwa alignments were combined for the subsequent chimeric read analysis using the RICpipe software. You filted the secondary alignment, but I have no idea about how to deal with the Supplementary Alignment of the alignment of bwa. Did you use option -M to flag shorter split hits as secondary?

Thank you.

https://github.com/alexdobin/STAR/archive/refs/tags/2.5.4b.zip

Hello,
Thanks for the code. But when I run the code of recalibrate_pvalue part, I got a mistake. Illegal division by zero at /public/data1/zhoumy/RIC/code/step5.screen_high-confidence_intermolecular/scripts/5.recalibrate_pvalue/2.multiple_testing_correction.pl line 244, line 1281814. The data is processed as described in your article. Can I delete this line? Or how should I handle this situation? Run the simulation again?

Thank you.

Hi,
Thank you for providing this nice pipeline for RIC data analysis.
I have some trouble in preparing gene annotation files. I use the reference hg38. How can I get the hg19.integrated.NCExon.bed.gz hg19.integrated.NCIntron.bed.gz hg19.integrated.PC3UTR.bed.gz hg19.integrated.PC5UTR.bed.gz hg19.integrated.PCCDS.bed.gz hg19.integrated.PCIntron.bed.gz? which is the meaning of PC?
Thank you!

Hi,
Thank you for providing this nice pipeline for RIC data analysis.
I met one issue while I tried to call intermolecular interaction.
In step: sh 3.calculate_pvalue/2.base_on_random/run.sh

The error messages are listed below:

Use of uninitialized value in addition (+) at /home/software/RICpipe-master/step5.screen_high-confidence_intermolecular/scripts/5.recalibrate_pvalue/merge_pvalue.pl line 40, line 24.
Illegal division by zero at /home/software/RICpipe-master/step5.screen_high-confidence_intermolecular/scripts/5.recalibrate_pvalue/merge_pvalue.pl line 45, line 24.
1 significant interactions
129079 other types interactions

Could you try to help? Thank you so much.

Y

Hi, Cao:

Thanks for your great work! I met one problem when running the script step1.collect_pair_tags/scripts/count_link_for_each_kind.pl, I cannot get the gapped number. After checking script, I found you define a subfunction to calculate gapped number:
sub part_count{
my $file=shift;
my $part_num;
open(IN,$file) || die;
while(my $line=){
chomp $line;
my @sub=split/\s+/,$line;
if($sub[0] =~ /Part_from_Align_Read/){
$part_num+=$sub[1];
}
}
close IN;
return $part_num;
}

This function aims to find the reads' name started with "Part_from_Align_Read" and sum the second column. But I have non reads which starts with "Part_from_Align_Read". The STAR vesion is 2.7.10b。 Only "Head" and "Tail" can be found at the beginning of reads' name. And I don't know what's the function of this subfunction, can it calculate the gapped reads number?
Looking forward to your reply

What is the rRNA sequence, reference, you use? Whether it is convenient for you to provide a download link