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BaM-seq and TBaM-seq, highly multiplexed and targeted RNA-seq protocols for rapid, low-cost library generation from bacterial samples [TBaM-seq]

ABSTRACT: Here, we report the development of two RNA-seq library preparation protocols that increase the throughput and decrease the cost of converting RNA to cDNA libraries compatible for sequencing on high-throughput platforms. BaM-seq allows for early barcoding of samples such that many biological samples can be processed simultaneously. TBaM-seq allows for enrichment of target RNAs to decrease the required sequencing depth. Both methods are able to accurately measure gene expression changes with high technical reproducibility and agreement with gold standard, lower throughput approaches.

ORGANISM(S): Bacillus subtilis

PROVIDER: GSE206424 | GEO | 2023/02/01

REPOSITORIES: GEO

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BaM-seq and TBaM-seq, highly multiplexed and targeted RNA-seq protocols for rapid, low-cost library generation from bacterial samples [BaM-seq]

Project description:Here, we report the development of two RNA-seq library preparation protocols that increase the throughput and decrease the cost of converting RNA to cDNA libraries compatible for sequencing on high-throughput platforms. BaM-seq allows for early barcoding of samples such that many biological samples can be processed simultaneously. TBaM-seq allows for enrichment of target RNAs to decrease the required sequencing depth. Both methods are able to accurately measure gene expression changes with high technical reproducibility and agreement with gold standard, lower throughput approaches.

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Transcription profiling of Drosophila ovaries from bam mutant and stwl bam double mutant females

Project description:Abstract: During Drosophila oogenesis, germline stem cell (GSC) identity is maintained largely by preventing the expression of factors that promote differentiation. This is accomplished via the activity of several genes acting either in the GSC or its niche. The translational repressors, Nanos and Pumilio, act in GSCs to prevent differentiation, likely by inhibiting translation of early differentiation factors, while niche signals prevent differentiation by silencing transcription of the differentiation factor Bam. We have found that the DNA-associated protein Stonewall (Stwl) is also required for GSC maintenance. stwl is required cell-autonomously; clones of stwl- germ cells were lost by differentiation, and ectopic Stwl caused an expansion of GSCs. stwl mutants acted as Suppressors of Variegation, indicating stwl normally acts in chromatin-dependent gene repression. In contrast to several previously described GSC maintenance factors, Stwl likely functions epigenetically to prevent GSC differentiation. Stwl-dependent transcriptional repression does not target bam, but rather Stwl represses the expression of many genes, including those that may be targeted by Nanos/Pumilio translational inhibition. Experiment Overall Design: Genetic variation comparison between germ cells of Drosophila ovaries from bam mutant and stwl bam double mutant females. Six total samples were analyzed, with each genotype performed in triplicates.

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