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Metode Perbanyakan Kontrol Positif Plasmodium spp. dengan Kloning
Positive control of Plasmodium spp. is needed in identification of malaria species using Polymerase Chain Reaction (PCR) method. The limitedness of malaria positive control encourages us to develop the cloning method to multiply Plasmodium spp. DNA particularly P.vivax, P. malariae, and P. ovale that are still unable to be cultured continuously. DNA product of single round and nested PCR were used as DNA target for cloning method.
The cloning method consisted of several processes which are ligation of DNA target into TOPO vector, transformation into competent Escherichia coli DH5α, and selection of transformation product using selective media. In the end of the process, PCR was done to check the success rate of cloning method.
Single round PCR resulting 200-300 bp of DNA product, while nested PCR resulting 100-200 bp of DNA product. The advantages of cloning method are produce large amount of products in such a shorter time in process with just small amounts of sample required. However, compared to continuous culture and blood specimen, DNA producted by cloning process can only be used for a spesific PCR method. Furthermore, other test is still required to examine stability of cloning product.
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