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Published on: May 16, 2013
Cloning of Plasmodium falciparum by single-cell sorting
Jun Miao1, Xiaolian Li, Liwang Cui
1Department of Entomology, The Pennsylvania State University, University Park, PA 16802, USA.
Experimental Parasitology
|May 4, 2010
Summary
This study introduces a rapid, single-cell sorting method for cloning malaria parasites, Plasmodium falciparum. This efficient technique improves parasite cloning accuracy and yield compared to traditional methods.
Area of Science:
- Malariology
- Parasitology
- Cell Biology
Background:
- Traditional malaria parasite cloning using limiting dilution is time-consuming and inaccurate.
- Limiting dilution cannot differentiate between single and multiple infected red blood cells (RBCs).
Purpose of the Study:
- To evaluate a single-cell sorting method for rapid cloning of Plasmodium falciparum.
- To improve the efficiency and accuracy of malaria parasite cloning.
Main Methods:
- Utilized a Plasmodium falciparum parasite engineered to express green fluorescent protein (GFP).
- Employed a cell sorter and a two-dimensional scattergram divided into 17 gates to isolate singly-infected erythrocytes.
- Sorted isolated cells into individual cultures for cloning.
Main Results:
- Achieved nearly 100% isolation of GFP-positive clones by pre-gating for GFP expression.
- Obtained a significantly higher number of parasite clones compared to the limiting dilution method.
- Molecular analyses confirmed the high homogeneity of parasite isolates obtained via single-cell sorting.
Conclusions:
- Single-cell sorting is a highly efficient method for cloning Plasmodium falciparum.
- This method offers advantages over traditional limiting dilution for both laboratory and clinical samples.
- The technique is valuable for cloning parasite populations from genetic manipulation and clinical studies.
