Figure 4
Expression and functional analyses of the PxABCH1 gene in P. xylostella.
(A) Expression fold changes of PxABCH1 in two Cry1Ac-resistant P. xylostella strains when compared to the susceptible P. xylostella strain. The expression levels of PxABCH1 were calculated based on the RPKM (the number of reads per kb of exon region per million mapped reads) values [log2ratio (DBM1Ac-R- or SZ-R-RPKM/DBM1Ac-S-RPKM)] of its unigenes derived from our previous transcriptome and RNA-Seq libraries38,39. (B) Relative expression levels of PxABCH1 as determined by qPCR in midgut tissues from fourth-instar larvae of all the susceptible and Cry1Ac/Btk-resistant P. xylostella strains. Expression of the RPL32 gene was used as the internal reference gene to normalize data sets and calculate expression levels. The relative expression levels (fold) were calculated by assigning an arbitrary value of 1 to the expression level in the susceptible DBM1Ac-S strain. Values shown in the figure are means and standard errors. Expression values with the same letter are not significantly different based on three biological replications and four technical repeats (P > 0.05; Holm-Sidak’s test; n = 3). (C) RNAi-mediated sublethal suppression of PxABCH1 expression after a low dose of its specific dsRNA (dsPxABCH1) injection at 48 h post-RNAi. The non-injection (control) or injection of P. xylostella larvae with buffer and dsEGFP were all used as negative controls. Expression of the RPL32 gene was used as the internal reference gene. The relative expression levels (fold) were calculated by assigning an arbitrary value of 1 to the expression level in the non-injection (control) DBM1Ac-S larvae. Values shown in the figure are means and standard errors. Different letters indicate significant differences between treatments (P < 0.05; Holm-Sidak’s test; n = 3). (D) Susceptibility of DBM1Ac-S larvae to two concentrations of Cry1Ac protoxin as affected by prior injection with buffer, dsEGFP or dsPxABCH1. Within each toxin concentration, the same letter indicate no significant difference between treatments (P > 0.05; Holm-Sidak’s test; n = 3).
