Estimating the proportion of resistance alleles from bulk Sanger sequencing, circumventing the variability of individual DNA

Masaaki Sudo; Kohji Yamamura; Shoji Sonoda; Takehiko Yamanaka

doi:10.1584/jpestics.D20-064

Estimating the proportion of resistance alleles from bulk Sanger sequencing, circumventing the variability of individual DNA

J Pestic Sci. 2021 May 20;46(2):160-167. doi: 10.1584/jpestics.D20-064.

Authors

Masaaki Sudo¹, Kohji Yamamura², Shoji Sonoda³, Takehiko Yamanaka²

Affiliations

¹ Institute of Fruit Tree and Tea Science, NARO, Kanaya Tea Research Station, 2769 Shishidoi, Kanaya, Shimada, Shizuoka 428-8501, Japan.
² Institute for Agro-Environmental Sciences, NARO, 3-1-3 Kannondai, Tsukuba, Ibaraki 305-8604, Japan.
³ School of Agriculture, Utsunomiya University, Utsunomiya, Tochigi 321-8505, Japan.

Abstract

Specimens should be examined as much as possible to obtain a precise estimate of the proportion of resistance alleles in agricultural fields. Monitoring traps that use semiochemicals on sticky sheets are helpful in this regard. However, insects captured by such traps are ordinarily left in the field until collection. Owing to DNA degradation, the amount of DNA greatly varies among insects, causing serious problems in obtaining maximum likelihood estimates and confidence intervals of the proportion of the resistance alleles. We propose a statistical procedure that can circumvent this degradation issue. R scripts for the calculation are provided for readers. We also propose the utilization of a Sanger sequencer. We demonstrate these procedures using field samples of diamide-resistant strains of the diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae). The validity of the assumptions used in the statistical analysis is examined using the same data.

Keywords: DNA degradation; Sanger sequencer; bulk sequencing; confidence interval; maximum likelihood estimation; proportion of resistance.