1. |
- Bagnoli, JW, et al.
(författare)
-
Sensitive and powerful single-cell RNA sequencing using mcSCRB-seq
- 2018
-
Ingår i: Nature communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 9:1, s. 2937-
-
Tidskriftsartikel (refereegranskat)abstract
- Single-cell RNA sequencing (scRNA-seq) has emerged as a central genome-wide method to characterize cellular identities and processes. Consequently, improving its sensitivity, flexibility, and cost-efficiency can advance many research questions. Among the flexible plate-based methods, single-cell RNA barcoding and sequencing (SCRB-seq) is highly sensitive and efficient. Here, we systematically evaluate experimental conditions of this protocol and find that adding polyethylene glycol considerably increases sensitivity by enhancing cDNA synthesis. Furthermore, using Terra polymerase increases efficiency due to a more even cDNA amplification that requires less sequencing of libraries. We combined these and other improvements to develop a scRNA-seq library protocol we call molecular crowding SCRB-seq (mcSCRB-seq), which we show to be one of the most sensitive, efficient, and flexible scRNA-seq methods to date.
|
|
2. |
|
|
3. |
|
|
4. |
|
|
5. |
- Janjic, A, et al.
(författare)
-
Prime-seq, efficient and powerful bulk RNA sequencing
- 2022
-
Ingår i: Genome biology. - : Springer Science and Business Media LLC. - 1474-760X. ; 23:1, s. 88-
-
Tidskriftsartikel (refereegranskat)abstract
- Cost-efficient library generation by early barcoding has been central in propelling single-cell RNA sequencing. Here, we optimize and validate prime-seq, an early barcoding bulk RNA-seq method. We show that it performs equivalently to TruSeq, a standard bulk RNA-seq method, but is fourfold more cost-efficient due to almost 50-fold cheaper library costs. We also validate a direct RNA isolation step, show that intronic reads are derived from RNA, and compare cost-efficiencies of available protocols. We conclude that prime-seq is currently one of the best options to set up an early barcoding bulk RNA-seq protocol from which many labs would profit.
|
|
6. |
|
|