Hi,

Agree with Guillem, edgeR and DESeq2 are the most stringent ones. Tuxedo package (TopHat-Bowtie) is very feasible too.

Good luck!

Depending on the number of replicates you have, you can use several packages. The most famous and used are edgeR and DESeq2, but if you are working without replicates, then you can try NOISeq, that helps to simulate replicates, to infer differential analysis on your samples.

Regards

Both edgeR and DESeq2 are really very good tools for DEG analysis and widely used. Depending on your data you can use anyone. DESeq2 gives more stringent and generally used for replicate data and for without replicating data it shows some warning message while running like "samples treated as replicate for estimation of dispersion". For detailed information, you can visit the website mentioned below. 

http://bioconductor.org/packages/devel/bioc/vignettes/DESeq2/inst/doc/DESeq2.html

And in the case of edgeR tool, it can be used for both replicate and without replicate data.

Thanks Giosa, Guillem, Rana, Rahul...

Rahul, DESeq2 is good but it is only working with data with replicates. I don't know how to run data without any replicates

Considering Domenico suggestion - I will try NOISeq for non replicate data to calculate DEG.

As u have data without replication so better your run both NOISeq and edgeR tool and compare both results. 

Hi Ellango

Some useful references 1

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=4&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFgg0MAM&url=http%3A%2F%2Fseqanswers.com%2Fforums%2Fshowthread.php%3Ft%3D23586&usg=AOvVaw0d9hr5geOW339Ka9Itgdry

2

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=5&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFgg8MAQ&url=https%3A%2F%2Fwww.biostars.org%2Fp%2F52535%2F&usg=AOvVaw0-ijdNR_G5NzSrq4M99na8

3

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=6&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFghCMAU&url=https%3A%2F%2Fwww.biostars.org%2Fp%2F94780%2F&usg=AOvVaw0LHuHEgZ0dX84yFKJgh02v

4

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=7&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFghJMAY&url=https%3A%2F%2Fwww.biostars.org%2Fp%2F119778%2F&usg=AOvVaw3UI97H6Dqvup5C5KBvy8je

5

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=8&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFghQMAc&url=http%3A%2F%2Fcole-trapnell-lab.github.io%2Fcufflinks%2Fcuffdiff%2F&usg=AOvVaw1pOWTawXjKhKSTezZ2AtI_

6

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=9&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFghWMAg&url=https%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fpmc%2Farticles%2FPMC4728800%2F&usg=AOvVaw1UsLXEwOR3x5XdDJLJIRzy

7

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=10&cad=rja&uact=8&ved=0ahUKEwiViY3erNnWAhVJKsAKHflIDV4QFghdMAk&url=http%3A%2F%2Fjournals.plos.org%2Fplosone%2Farticle%3Fid%3D10.1371%2Fjournal.pone.0103207&usg=AOvVaw0RDEwyE0EBAWtgGKgYobCM

8

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=11&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCCYwAA&url=https%3A%2F%2Fsupport.bioconductor.org%2Fp%2F63643%2F&usg=AOvVaw3eCLt3qdOKjFd-NDCRreae

9

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=12&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCCwwAQ&url=https%3A%2F%2Fbiostar.usegalaxy.org%2Fp%2F5336%2F&usg=AOvVaw3v16uPDOviob4VlkBvolzJ

10

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=13&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCDIwAg&url=http%3A%2F%2Fjournals.plos.org%2Fplosone%2Farticle%3Fid%3D10.1371%2Fjournal.pone.0103207&usg=AOvVaw0RDEwyE0EBAWtgGKgYobCM

11

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=14&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCDwwAw&url=http%3A%2F%2Fsoftware.broadinstitute.org%2Fcancer%2Fsoftware%2Fgenepattern%2Fmodules%2Fdocs%2FCuffdiff%2F7&usg=AOvVaw0IUE0Rfn5tSVn2sEFzYlY9

12

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=15&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCEEwBA&url=https%3A%2F%2Fbmcbioinformatics.biomedcentral.com%2Farticles%2F10.1186%2F1471-2105-14-91&usg=AOvVaw34MeoGlf4C3PTlj09MiHIE

13

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=16&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCEswBQ&url=http%3A%2F%2Fwww.mdpi.com%2F2079-6374%2F3%2F3%2F238%2Fpdf&usg=AOvVaw0i_z0HHftMuWmzVzibV7pX

Q

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=17&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCFUwBg&url=http%3A%2F%2Fwww.cureffi.org%2F2013%2F09%2F12%2Fcounts-vs-fpkms-in-rna-seq%2F&usg=AOvVaw0kZxqbDJYVXxXbPxvVXrQX

w

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=18&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCFwwBw&url=https%3A%2F%2Fsupport.illumina.com%2Fcontent%2Fdam%2Fillumina-support%2Fdocuments%2Fdocumentation%2Fsoftware_documentation%2Fbasespace%2Fbasespace-cufflinks-assembly-v2-app-guide-1000000006108-00.pdf&usg=AOvVaw3rQpnhCH2blwRSwStSzHSU

e

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=19&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCGEwCA&url=https%3A%2F%2Fen.wikipedia.org%2Fwiki%2FList_of_RNA-Seq_bioinformatics_tools&usg=AOvVaw3lrtWRFrfo7rmY8vJIE7O4

r

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=20&cad=rja&uact=8&ved=0ahUKEwigu8_JrtnWAhXJ2RoKHZo2Be04ChAWCGcwCQ&url=https%3A%2F%2Fwww.melbournebioinformatics.org.au%2Ftutorials%2Ftutorials%2Frna_seq_dge_advanced%2Frna_seq_advanced_tutorial%2F&usg=AOvVaw2VWPFANDF4o9LHmgMEJsQl

y

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=21&cad=rja&uact=8&ved=0ahUKEwi9sqOgr9nWAhVCPRoKHXoGCCA4FBAWCCYwAA&url=https%3A%2F%2Fpods.iplantcollaborative.org%2Fwiki%2Fdisplay%2FTUT%2FRNA-seq%2BTutorial-%2BHISAT2%252C%2BStringTie%2Band%2BBallgown&usg=AOvVaw34BqbBqhZmm0FXEkFuG-5c

t

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=22&cad=rja&uact=8&ved=0ahUKEwi9sqOgr9nWAhVCPRoKHXoGCCA4FBAWCCkwAQ&url=https%3A%2F%2Fdocs.uabgrid.uab.edu%2Fwiki%2FUAB_Galaxy_RNA_Seq_Step_by_Step_Tutorial&usg=AOvVaw1zWsNceQuD0KbXFGoOc3LO

u

https://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=23&cad=rja&uact=8&ved=0ahUKEwi9sqOgr9nWAhVCPRoKHXoGCCA4FBAWCDAwAg&url=https%3A%2F%2Fdash.harvard.edu%2Fbitstream%2Fhandle%2F1%2F29407854%2F5002174.pdf%3Fsequence%3D1&usg=AOvVaw0CROZzVcg_tQo0D6f26Tw1

Thanks Laith, Rahul