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MRNA sequence analysis of zebrafish embryos injected with the codon tag reporter library

ABSTRACT: mRNA sequence analysis of zebrafish embryos injected with the codon tag reporter library

PROVIDER: PRJNA724039 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14308848.fastq.gz	Fastqsanger.gz
	SRR14308849.fastq.gz	Fastqsanger.gz
	SRR14308850.fastq.gz	Fastqsanger.gz
	SRR14308851.fastq.gz	Fastqsanger.gz
	SRR14308852.fastq.gz	Fastqsanger.gz

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Project description:A ELMSeq reporter cassette was created to monitor Dam levels by methylation, and introduced in the genome. The regions of 6 nt upstream and 6 nt downstream the stop codon were randomized to study their effect on gene expression. The ELMSeq reporter cassette was composed of: promoter - dam - random N6 - stop codon TAA - random N6 - spacer - 4xGATC. The amplicon was spanning the C-terminal region. The cassette was introduced in Mycoplasma pneumoniae.

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ENVIRONMENT MODULATES PROTEIN HETEROGENEITY THROUGH TRANSCRIPTIONAL AND TRANSLATIONAL STOP CODON RECODING

Project description:In order to systematically assess the frequency and origin of stop codon recoding events, we designed a library of reporters. We introduced premature stop codons into mScarlet that enabled high-throughput quantification of protein synthesis termination errors in E. coli using fluorescent microscopy. We found that under stress conditions, stop codon recoding may occur with a rate as high as 80%, depending on the nucleotide context, suggesting that evolution frequently samples stop codon recoding events. The analysis of selected reporters by mass spectrometry and RNA-seq showed that not only translation but also transcription errors contribute to stop codon recoding. The RNA polymerase is more likely to misincorporate a nucleotide at premature stop codons. Proteome-wide detection of stop codon recoding by mass spectrometry revealed that temperature regulates the expression of cryptic sequences generated by stop codon recoding in E. coli. Overall, our findings suggest that the environment influences the accuracy of protein production which increases protein heterogeneity when the organisms need to adapt to new conditions.

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