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SUPPLEMENTARY MATERIAL – Parr JB et al. Malar J 2018.

https://doi.org/10.1186/s12936-018-2287-4

1

SUPPLEMENTARY MATERIAL

Figure S1 – Pfhrp2 assay performance using serially diluted P. falciparum 3D7 strain DNA. Elongation

temperatures were varied as listed below. All other reaction conditions are specified in Table 1.

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SUPPLEMENTARY MATERIAL – Parr JB et al. Malar J 2018.

https://doi.org/10.1186/s12936-018-2287-4

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Figure S2 – Pfhrp3 assay performance using serially diluted P. falciparum 3D7 strain DNA. Elongation

temperatures were varied as listed below. All other reaction conditions are specified in Table 1.

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SUPPLEMENTARY MATERIAL – Parr JB et al. Malar J 2018.

https://doi.org/10.1186/s12936-018-2287-4

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Figure S3 – Representative agarose gel electrophoresis depicting unexpected spurious bands from Dd2

strain (pfhrp2-deleted) control DNA. PCR targeting pfhrp2 exon 1/2 (assay 1 outer) yielded a spurious

~300bp band from serial dilutions of pfhrp2-deleted Dd2 strain control DNA at all three elongation

temperatures.

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SUPPLEMENTARY MATERIAL – Parr JB et al. Malar J 2018.

https://doi.org/10.1186/s12936-018-2287-4

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Figure S4 – Representative agarose gel electrophoresis depicting unexpected spurious bands from HB3

strain (pfhrp3-deleted) control DNA. PCR targeting pfhrp3 exon 1/2 (assay 5) yielded spurious bands at

~300, 400, and 800bp from serial dilutions of Dd2 strain control DNA using optimized elongation

temperatures (Table 1).

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SUPPLEMENTARY MATERIAL – Parr JB et al. Malar J 2018.

https://doi.org/10.1186/s12936-018-2287-4

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1 REF: pfhrp2 A T G G T T T C C T T C T C A A A A A A T A A A G T A T T A T C C G C T G C C G T T T T T G C C T C C G T A C T T T T G T T A G A T A A C g t a a g c a t t REF: pfhrp3 . . . A . . . T . . . t . . . 3D7_hrp2exon1/2 . . . . DD2_hrp2exon1/2 . . . t . . . 79 REF: pfhrp2 t t a a t t g c a a a t a g a a a t a a a t a t a t c a t t c a a t t a t a a a a a t a c a t a t a a t a a t a a t a t a t a t a t a t a t a t a t a t a t REF: pfhrp3 . . . t . . . a . . c . t . c . . . t . . . . 3D7_hrp2exon1/2 . . . . DD2_hrp2exon1/2 . . . t . . . a . . c . t . c . . . t . . . . 157 REF: pfhrp2 t t a t a t a t - - - t g t a t a t a t a t a a a t t t t t t c a t t t t t a a a t g c t t t t t t a t t t t t a t a t a g A A T A A T T C C G C A T T REF: pfhrp3 a . . . a t a t a . a . . . g . . c . . . a . . . a . . . c . . . - - . . . C . . . . A . . . 3D7_hrp2exon1/2 . . . - - - . . . . DD2_hrp2exon1/2 a . . . - - - . . . g . . c . . . a . . . a . . . c . . . - - . . .

Figure S5 – The sequence of pfhrp2 exon 1/2 (assay 1) PCR product aligns to pfhrp3, due to spurious PCR amplification of the Dd2 pfhrp3

gene. PCR was performed using 0.01ng/μL of 3D7 (pfhrp2-positive) and Dd2 (pfhrp2-negative) control DNA, respectively (see Figure S3),

followed by Sanger sequencing of amplicons. Reference sequences from the consensus 3D7 (v3.0) genome for pfhrp2 and pfhrp3 are

displayed on the top two rows (REF), from 5’->3’, with capital letters for coding regions and genetic coordinates in reference to the pfhrp2

gene. Identical bases are indicated by a period (.), missing bases by a dash (-), substitutions by the discordant base. PCR product sequence

contigs are highlighted as follows: 3D7 control DNA (light gray); Dd2 control DNA (dark gray).

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SUPPLEMENTARY MATERIAL – Parr JB et al. Malar J 2018.

https://doi.org/10.1186/s12936-018-2287-4

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Figure S6 – The sequences of pfhrp3 exon 1/2 (assay 5) PCR product align to pfhrp2, due to spurious PCR amplification of the HB3 pfhrp2

gene. PCR was performed using 0.01ng/μL of 3D7 (pfhrp3-positive) and HB3 (pfhrp3-negative) control DNA, respectively (see Figure S4),

followed by Sanger sequencing of amplicons. Reference sequences from the consensus 3D7 (v3.0) genome for pfhrp2 and pfhrp3 are

displayed on the top two rows (REF), from 5’->3’, with capital letters for coding regions and genetic coordinates in reference to the pfhrp3

gene. Identical bases are indicated by a period (.), missing bases by a dash (-), substitutions by the discordant base. PCR product sequence

contigs are highlighted as follows: 3D7 control DNA (medium gray) and HB3 control DNA 300bp fragment (light gray), 400bp fragment

(medium gray), and 800bp fragment (dark gray).

1 REF: pfhrp3 A T G G T T T C C T T C T C A A A A A A T A A A A T A T T A T C C G C T G C C G T T T T T G C T T C C G T A C T T T T G T T A G A T A A C g t a a g t a t t REF: pfhrp2 . . . G . . . C . . . c . . . 3D7_hrp3exon1/2 HB3_hrp3exon1/2_300bp HB3_hrp3exon1/2_400bp HB3_hrp3exon1/2_800bp 79 REF: pfhrp3 t t a a t t g c a a a t a t a a a t a a a t a t a a c a c t t a c t t a t a a a a a t a c a t a t a a t a a t a t t a t a t a t a t a t a t a t a t a t a t REF: pfhrp2 . . . g . . . t . . t . c . a . . . a . . . . 3D7_hrp3exon1/2 . . . - - . . . . HB3_hrp3exon1/2_300bp . . . t . . t . c . a . . . a . . . . HB3_hrp3exon1/2_400bp HB3_hrp3exon1/2_800bp . . . g . . . t . . t . c . a . . . a . . . . 157 REF: pfhrp3 a t a t a t a t a t a t a t a t a t a t a t g t a c a t t t t t a c a t t t t t a a a t g a t t t t t c a t t t t t a t a - - g A A T A A C T C C G A A T T REF: pfhrp2 t . . . - - - . g . . . a . . a . . . t . . . c . . . t . . . t a . . . T . . . . C . . . 3D7_hrp3exon1/2 . . . . HB3_hrp3exon1/2_300bp . . . . HB3_hrp3exon1/2_400bp HB3_hrp3exon1/2_800bp . . . . t . . . - . g . . . a . . a . . . t . . . c . . . t . . . t a 233 REF: pfhrp3 T A A C A A T A A C T T G T T T A G C A A A A A T G C A A A A G G A C T T A A T T C A A A T A A G A G A T T A T T A C A C G A A A G T C A A G C A C A T G C REF: pfhrp2 . . . T . . . G . . . T . . . C . . . T 3D7_hrp3exon1/2 . . . . HB3_hrp3exon1/2_300bp HB3_hrp3exon1/2_400bp . . . . T HB3_hrp3exon1/2_800bp 311 REF: pfhrp3 A G G T G A T G C C C A T C A T G C A C A T C A T G T A G C T G A T G C C C A T C A T G C A C A T C A T G T A G C T G A T G C C C A T C A T G C A C A T C A REF: pfhrp2 . . A . . . T . . . C . . . T . . . C . . . T . . . . . 3D7_hrp3exon1/2 HB3_hrp3exon1/2_300bp HB3_hrp3exon1/2_400bp . . A . . . T . . . C . . . T . . . C . . . T . . . . . HB3_hrp3exon1/2_800bp 389 REF: pfhrp3 T G T A G C T G A T G C C C A T C A T G C A C A T C A T G T A G C T G A T G C C C A T C A T G C A C A T C A T G C T C A C C A T G C A G C T A A T G C T C A REF: pfhrp2 . . . C . . . T . . . C . . . C . . . T . . . A G C . G . . . . C C A . C . . . . 3D7_hrp3exon1/2 HB3_hrp3exon1/2_300bp HB3_hrp3exon1/2_400bp . . . C . . . T . . . C . . . . HB3_hrp3exon1/2_800bp

References

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