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Fig. 4 | Parasites & Vectors

Fig. 4

From: Molecular characterization and genetic authentication assay for Anopheles ‘hemocyte-like’ cell lines 4a-3A and 4a-3B

Fig. 4

Molecular assay to differentiate 4a-3A, 4a-3B, Ag55 and Sua4.0 cell lines. A Molecular fingerprints of three PCRs amplifying indel regions in 4a-3A (lanes 2–4), 4a-3B (lanes 5–7), Ag55 (lanes 8–10), Sua4.0 cells (lanes 11–13) and no PCR template controls (lanes 14–16); 100-bp ladder (lane 1), indel 2R.25670547 (lanes 2, 5, 8, 11, 14), indel 3R.11788474 (lanes 3, 6, 9, 12, 15), indel 3R.11809836 (lanes 4, 7, 10, 13, 16) and 1-kb Plus DNA Ladder (lane 17). B Table of expected PCR product sizes for 2R.25670547, 3R.11788474 and 3R.11809836 for each of the cell lines based on Sanger sequencing results. *Predicted size based on whole genome sequencing was 532 bp for 3R.11788474 in 4a-3A genomic DNA; however, upon Sanger sequencing an additional indel of 204-bp size was discovered following the predicted 148-bp indel resulting in an amplicon size of 736 bp. All sizes were verified by Sanger sequencing. C Detection of contamination by indel assay. The 3R.11788474 indel assay is able to detect contaminating genomic DNA present at 10% of the total PCR template input. Lane 1 GeneRuler 1-kb Plus DNA Ladder; lane 2: 20 ng 4a-3B gDNA; lane 3: 18 ng 4a-3B gDNA and 2 ng 4a-3A gDNA; lane 4: 16 ng 4a-3B gDNA and 4 ng 4a-3A gDNA; lane 5: 13.3 ng 4a-3B gDNA and 6.6 ng 4a-3A gDNA; lane 6: 10 ng 4a-3B gDNA and 10 ng 4a-3A gDNA; lane 7: 6.6 ng 4a-3B gDNA and 13.3 ng 4a-3A gDNA; lane 8: 4 ng 4a-3B gDNA and 16 ng 4a-3A gDNA; lane 9: 2 ng 4a-3B gDNA and 18 ng 4a-3A; lane 10: 20 ng 4a-3A gDNA, lane 11: control, no PCR template, lane 12: 100-bp ladder

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