Strand-specific libraries for high throughput RNA sequencing (RNA-Seq) prepared without poly(A) selection

Silence

Table 1 Comparison of library construction protocols

Method	Time (days)	Total steps	Reagent cost ($)	Starting RNA (ng)	Use for small RNA	Kits available	Barcoding described
This protocol	2	9	38	50	No	No	Yes
RNA ligation	8	19	250	1200	Yes	No	No
Illumina RNA ligation	5	16	240	100	Yes	Partially	No
Illumina RNA ligation-SPRI	4	12	220	100	Yes	Partially	No
SMART	5	8	80	100	Yes	No	No
Hybrid	5	13	90	500	Yes	No	No
NNSR	4	9	90	500	Unclear	No	No
NNSR, no Actinomycin D	4	9	90	500	Unclear	No	No
Bisulfite ‘S’	6	19	540	1000	No	Mostly	No
Bisulfite ‘H’	6	19	540	1000	No	Mostly	No
dUTP	5	17	430	200	No	Mostly	No
dUTP oligo(dT)	5	17	440	200	No	Mostly	No

Data for other methods from Levin [12]. Values only include steps after rRNA removal (this protocol) or poly(A) selection. dUTP: deoxyuridine triphosphate; NNSR: not not so random; SMART: switching mechanism at 5’ end of RNA transcript; SPRI, solid phase reversible immobilization.