Sam2TPM_RPKM

inputs: sample.sam, sample.gtf

Usage:

1. place py in a directory with your sample.sam and sample.gtf
2. run with python (i.e $python Sam2tpm_rpkm.py)

How it works:

###Calculate TPM:

(X[i]/L[i])*(1/(sum(Xtot/Ltot)))*1e6

####breaking it down to steps:

1. Take each gene read and divide by length (in kb) of gene
2. Next take these normalized reads and sum them for each gene
3. create scaling factor by dividing summed normalized reads by 1e6
4. take your normalized reads from step 1 and divide by the scaling factor

####Taken from these websites: Instructions from [here] (https://www.youtube.com/watch?v=TTUrtCY2k-w) and [here] (https://haroldpimentel.wordpress.com/2014/05/08/what-the-fpkm-a-review-rna-seq-expression-units/)

###Calculate RPKM:

Mapped_Reads/((Total_reads/1000000) * (Lenth_Gene/1000))

#####Mapped_Reads is the reads that were mapped to a gene in the gtf file #####Total_reads is the total reads in the experiment #####Length of gene is in kb

1. Get number of reads
2. Calculate Number_of_reads/(length_of_gene/kilobase * total_number_reads/1e6)

Instructions found here

##How to use:

Run it in the command line as such:

$python SamToTpm.py

Input: Input sam file must be named: "sample.sam" Input gtf file must be named: "sample.gtc"

Example sam:

@HD	VN:1.3	SO:coordinate
@SQ	SN:reference	LN:5528445
HWI-ST1426:114:HGTH7ADXX:1:1202:2398:51916	163	reference	4	60	101M	=	37	134	TTTTCATTCTGACTGCAACGGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAGCTTCTGAACTGGTTACCTGCCGTGAGTAAATTAAA	BBBFFFFFFFFFFIIIIIIIIIIIIIIIFIFIIIIFFFFIIIFFIIIIIIIIIIIIIIFFFFFFFFFFFFFFFFBFFFFFFFFFFFFFBFFF<BBBFFFFF	XT:A:U	NM:i:0	SM:i:37	AM:i:37	X0:i:1	X1:i:0	XM:i:0	XO:i:0	XG:i:0	MD:Z:101
HWI-ST1426:114:HGTH7ADXX:1:1216:11691:62785	99	reference	11	29	35S24M42S	=	35	101	GGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACTGCAACGGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAGCTTCTGA	BBBFFFFFFBBFBFFFFFFFFFFFBF<BFIFFFBFIFFFFFFIFIFIIII<FFFFIIFBFFFFBFBFFFBFBBB<77BBBFFBBBBBFBFF<BBFFBBBBB	XT:A:M	NM:i:0	SM:i:29	AM:i:29	XM:i:0	XO:i:0	XG:i:0	MD:Z:24
HWI-ST1426:114:HGTH7ADXX:1:2203:7459:45734	117	reference	16	0	*	=	16	0	TCAGAAGCTGCTGTCAGAGACTCTTTTTTTAATCCACACAGAGACATATTGCCCGTTGCAGTCAGAGACTCTTTTTTTAATCCACACAGAGACATATTGCC	BB<<FB<BBBBBBBBFFBBBBB<7BB<BBBBB<FFBBBBB7<<<FFIIIIIFBFFFFBFIIFBFBFFFIIIFFBFIFFFFBFFFFFIIFFBFFFFFFFBBB
HWI-ST1426:114:HGTH7ADXX:1:2203:7459:45734	153	reference	16	37	54M	=	16	0	CTGCAACGGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAG	BB<BBB<BB<<BBBBBBBBB<<BFBBBBFB<FBFIIFFFFFFFFIFFFFFFB77	XT:A:U	NM:i:0	SM:i:37	AM:i:0	X0:i:1	X1:i:0	XM:i:0	XO:i:0	XG:i:0	MD:Z:54
HWI-ST1426:114:HGTH7ADXX:1:1203:18673:44805	99	reference	22	60	101M	=	126	205	CGGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAGCTTCTGAACTGGTTACCTGCCGTGAGTAAATTAAAATTTTATTGACTTAGGTC	BBBFFFFFFFFFFFIIIIIFFIBFFIFFIIIIIIIIIIIFFFIIIIIFIFFIIIBBFFFFIIIFFFFFFFFFBBBBBFFFBBFFFFFFFFFBBFBFFFFBB	XT:A:U	NM:i:0	SM:i:37	AM:i:37	X0:i:1	X1:i:0	XM:i:0	XO:i:0	XG:i:0	MD:Z:101
HWI-ST1426:114:HGTH7ADXX:1:2212:5218:9001	163	reference	22	60	101M	=	126	205	CGGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAGCTTCTGAACTGGTTACCTGCCGTGAGTAAATTAAAATTTTATTGACTTAGGTC	BBBFFFFFFFFFFIIIIFIIIIIIIIIIIIIIIIIFIIIIIIIIIIIIIIIIIIIIIIIIFIIFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFBFFFBB	XT:A:U	NM:i:0	SM:i:37	AM:i:37	X0:i:1	X1:i:0	XM:i:0	XO:i:0	XG:i:0	MD:Z:101
HWI-ST1426:114:HGTH7ADXX:1:1107:18270:12891	163	reference	23	60	101M	=	73	151	GGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAGCTTCTGAACTGGTTACCTGCCGTGAGTAAATTAAAATTTTATTGACTTAGGTCA	BBBFFFFFFFFFFIIIFIFIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF	XT:A:U	NM:i:0	SM:i:37	AM:i:37	X0:i:1	X1:i:0	XM:i:0	XO:i:0	XG:i:0	MD:Z:101
HWI-ST1426:114:HGTH7ADXX:1:1211:11146:4928	99	reference	23	60	101M	=	73	151	GGGCAATATGTCTCTGTGTGGATTAAAAAAAGAGTCTCTGACAGCAGCTTCTGAACTGGTTACCTGCCGTGAGTAAATTAAAATTTTATTGACTTAGGTCA	BBBFFFFFFFFFFIIIFIFFFIIIIIIIIIIIIIFFFIIIIIIIIIFIIIIIIIIIIIIFIFIIIFFFFFFFFFBFFFFFFFBFFBFFFFBFFFFFFFBFF	XT:A:U	NM:i:0	SM:i:37	AM:i:37	X0:i:1	X1:i:0	XM:i:0	XO:i:0	XG:i:0	MD:Z:101

sample gtc file:

reference	CDS	10	104	.	+	1	something
reference	CDS	114	281	.	+	1	something

Output: The output will come in the command line:

Example:

Gene: 'reference', Feature: 'CDS', TPM: 42711772.175, RPKM: 2417794.971
Gene: 'reference', Feature: 'CDS', TPM: 91118447.306, RPKM: 2903284.34