Question

Error in stattest function

0

Entering edit mode

amy16 • 0

@amy16-14425

Last seen 6.2 years ago

Hi all,

Please let me know how I should go about in fixing this error.

> pData(bg)
     ids  treatment  genotype
1 B1C_01    control resistant
2 B1T_01 inoculated resistant
> prr01_trans_result=stattest(bg_filt, feature="transcript", covariate="treatment", getFC=TRUE, meas="FPKM")
Error in stattest(bg_filt, feature = "transcript", covariate = "treatment",  :
  There must be at least two replicates per group. Make sure covariate is categorical; if continuous, consider the timecourse option, or specify your own models with mod and mod0.

PS: I do not have replicates for this experiment.

Thanks

R rnaseq Ballgown • 2.7k views

ADD COMMENT • link updated 3.5 years ago by muhammadharoon974 • 0 • written 6.9 years ago by amy16 • 0

score 1 · Answer 1 · 2018-01-29

1

Entering edit mode

Alyssa Frazee ▴ 210

@alyssa-frazee-6710

Last seen 4.1 years ago

San Francisco, CA, USA

The error message is there because it is not possible to use ballgown's stattest function when you only have one sample per condition. You cannot perform statistical inference without replicates. However, all is not lost! If you want, you can rank the genes/transcripts by fold change, for example, by calculating log2(FPKM_rep1) - log2(FPKM_rep2), and you may be able to get some useful information that way. But without replicates, you can not perform statistical inference, so you can't calculate p-values, confidence intervals, standard errors or anything like that. (Variance is undefined when you have only one item). This is true for Ballgown and any other software you might use for this problem.

ADD COMMENT • link 6.9 years ago Alyssa Frazee ▴ 210

0

Entering edit mode

Thanks Alyssa for the reply and the good news that I can rank the genes/transcripts by fold change. Is this fold change calculation script described in the Ballgown manual? Can u please direct me on how I go about it?

PS: I understand and I totally agree with you that without replicates, one can not perform statistical inference, so can't calculate p-values, confidence intervals, standard errors or anything like that. (Variance is undefined when you have only one item).

Thanks

Amy

ADD REPLY • link 6.9 years ago amy16 • 0

0

Entering edit mode

Please see https://github.com/alyssafrazee/ballgown#accessing-assembly-data for information on how to access the fold change information for each transcript (or gene or exon) for each sample.

You will then need to write your own R code to calculate the expression fold change between the two samples for each transcript.

Good luck!

ADD REPLY • link 6.9 years ago Alyssa Frazee ▴ 210

0

Entering edit mode

Thanks for the reply...

ADD REPLY • link 6.9 years ago • updated 6.2 years ago amy16 • 0

score 0 · Answer 2 · 2021-05-27

0

Entering edit mode

muhammadharoon974 • 0

@dfb28f64

Last seen 3.5 years ago

Pakistan

Hi, I have only three samples and there is no replicate. I am facing the same problem. Can you share your script. Thanks!

ADD COMMENT • link 3.5 years ago muhammadharoon974 • 0