# How I deal with this kind of gene expression comparision

I have RNA-seq from two sequencing batches; Lab technician says that he has run the RNA expression quantification two times in bathes 1 and 2 for example tumor 1 in batch 1 and tumor 1 in batch 2 , normal 2 in batch1 and normal 2 in batch 2. PCA shows however shows two experimental runs are too close to each other and can be merged

[![enter image description here][1]][1]

Now my question is can I compare only a tumour sample with its own matched normal sample in one patient to get differentially expressed genes for this patient by using experimental runs (technical replicates )? Design would be Tumour1_batch1 Tumour1_batch2 versus Normal1_batch1 Normal1_bacth2

> head(mycols)
Condition
Normal1_batch1  N
Normal1_batch2  N
Tumour1_batch1  T
Tumour1_batch2  T

• Yes, you could (although I think you would need three samples in each side to be able to calculate the sd and var). However, how generalizable it would be ? What is your biological question behind this comparison? One thing is to pair the samples and control for batch effect and another is to just compare 4 samples. – llrs Sep 17 '19 at 12:54
• My main question here is: what are the differentially expressed genes between tumour and normal adjacent sample for this specific patient where I am going to use two experimental runs as my replications for having statistical power . Thank you – Exhausted Sep 17 '19 at 12:57
• Using technical replicates tells you how good your technician can use a pipette, it gives you no information about the biology behind it. In statistics we want to sample (take samples from a population) to get info about the whole population. For example, if you want to have the average length from a population, you sample different people to get a good average. In your case you would measure one person twice, it tells you nothing about the population but it tells how good your measuring skills are. – benn Sep 17 '19 at 13:48
• If you want to compare individual patients, why are there less normal samples compared to tumor samples? – PPK Sep 18 '19 at 13:51
• Because I don't have real replication rather two experimental runs – Exhausted Sep 19 '19 at 11:05