A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites
Abstract Background The transcriptional regulation that occurs in malaria parasites during the erythrocytic stages of infection can be studied in vivo with rodent malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific...
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| Format: | Article |
| Language: | English |
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BMC
2019-01-01
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| Series: | Malaria Journal |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s12936-019-2659-4 |
| _version_ | 1818996012094586880 |
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| author | Abhinay Ramaprasad Amit Kumar Subudhi Richard Culleton Arnab Pain |
| author_facet | Abhinay Ramaprasad Amit Kumar Subudhi Richard Culleton Arnab Pain |
| author_sort | Abhinay Ramaprasad |
| collection | DOAJ |
| description | Abstract Background The transcriptional regulation that occurs in malaria parasites during the erythrocytic stages of infection can be studied in vivo with rodent malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. Results A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the rodent malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in malaria parasites. Conclusions Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in rodent malaria parasites. |
| first_indexed | 2024-12-20T21:22:58Z |
| format | Article |
| id | doaj.art-76ee734b6c2e43039524fbe5e0e76d9e |
| institution | Directory Open Access Journal |
| issn | 1475-2875 |
| language | English |
| last_indexed | 2024-12-20T21:22:58Z |
| publishDate | 2019-01-01 |
| publisher | BMC |
| record_format | Article |
| series | Malaria Journal |
| spelling | doaj.art-76ee734b6c2e43039524fbe5e0e76d9e2022-12-21T19:26:13ZengBMCMalaria Journal1475-28752019-01-0118111110.1186/s12936-019-2659-4A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasitesAbhinay Ramaprasad0Amit Kumar Subudhi1Richard Culleton2Arnab Pain3Pathogen Genomics Laboratory, Biological and Environmental Sciences and Engineering (BESE) Division, King Abdullah University of Science and Technology (KAUST)Pathogen Genomics Laboratory, Biological and Environmental Sciences and Engineering (BESE) Division, King Abdullah University of Science and Technology (KAUST)Malaria Unit, Department of Pathology, Institute of Tropical Medicine (NEKKEN), Nagasaki UniversityPathogen Genomics Laboratory, Biological and Environmental Sciences and Engineering (BESE) Division, King Abdullah University of Science and Technology (KAUST)Abstract Background The transcriptional regulation that occurs in malaria parasites during the erythrocytic stages of infection can be studied in vivo with rodent malaria parasites propagated in mice. Time-series transcriptome profiling commonly involves the euthanasia of groups of mice at specific time points followed by the extraction of parasite RNA from whole blood samples. Current methodologies for parasite RNA extraction involve several steps and when multiple time points are profiled, these protocols are laborious, time-consuming, and require the euthanization of large cohorts of mice. Results A simplified protocol has been designed for parasite RNA extraction from blood volumes as low as 20 μL (microsamples), serially bled from mice via tail snips and directly lysed with TRIzol reagent. Gene expression data derived from microsampling using RNA-seq were closely matched to those derived from larger volumes of leucocyte-depleted and saponin-treated blood obtained from euthanized mice with high reproducibility between biological replicates. Transcriptome profiling of microsamples taken at different time points during the intra-erythrocytic developmental cycle of the rodent malaria parasite Plasmodium vinckei revealed the transcriptional cascade commonly observed in malaria parasites. Conclusions Microsampling is a quick, robust and cost-efficient approach to sample collection for in vivo time-series transcriptomic studies in rodent malaria parasites.http://link.springer.com/article/10.1186/s12936-019-2659-4Rodent malaria parasitesTranscriptomicsTime-seriesPlasmodiumMalariaMicrosampling |
| spellingShingle | Abhinay Ramaprasad Amit Kumar Subudhi Richard Culleton Arnab Pain A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites Malaria Journal Rodent malaria parasites Transcriptomics Time-series Plasmodium Malaria Microsampling |
| title | A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites |
| title_full | A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites |
| title_fullStr | A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites |
| title_full_unstemmed | A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites |
| title_short | A fast and cost-effective microsampling protocol incorporating reduced animal usage for time-series transcriptomics in rodent malaria parasites |
| title_sort | fast and cost effective microsampling protocol incorporating reduced animal usage for time series transcriptomics in rodent malaria parasites |
| topic | Rodent malaria parasites Transcriptomics Time-series Plasmodium Malaria Microsampling |
| url | http://link.springer.com/article/10.1186/s12936-019-2659-4 |
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