Comparison of RNAi Sequences in Insect-Resistant Plants to Expressed Sequences of a Beneficial Lady Beetle: A Closer Look at Off-Target Considerations

Abstract

Sequences obtained from transcriptomes of the lady beetle Coleomegilla maculata were compared to those designed for incorporation into crops. Searches of the transcriptomes identified sequences as the most likely to be closely similar to the sequences described in RNAi plant incorporated products. Some proposed prime RNAi pest management targets were also used to identify predicted orthologs from C. maculata. The lady beetle sequences were aligned with sequences from corn rootworms and Colorado potato beetles and, as appropriate in the case of targets, regions of similarity were compared with the genetic model organism for beetles, Tribolium castaneum. Some high levels of nucleotide identity were identified, particularly with an actin-derived sequence from Colorado potato beetle. This actin-derived sequence shared identical sequences with the lady beetle and a parasitic wasp.

Keywords: beneficial organism; genetic pest control; lady beetle; risk assessment.

Figure 1

Predicted sequence and translation from Coleomegilla maculata transcriptomes most similar to Diabrotica virgifera virgifera Snf7 used to design rootworm resistant maize. (A) Complete cDNA sequence. Highlighted section is the portion of the sequence matching the maize RNAi transgene. Upper case indicates translated sequence, lower case untranslated; (B) Alignment of the RNAi transgene regions. Lines between nucleotides indicate identity, identical letters between amino acid sequences indicate identity. The symbol (+) indicates an amino acid substitution by a similar residue, while a (0) or a blank between nucleotide or amino acid letter, respectively, indicates a non-similar substitution. Bottom: amino acid alignment.

Figure 1

Predicted sequence and translation from Coleomegilla maculata transcriptomes most similar to Diabrotica virgifera virgifera Snf7 used to design rootworm resistant maize. (A) Complete cDNA sequence. Highlighted section is the portion of the sequence matching the maize RNAi transgene. Upper case indicates translated sequence, lower case untranslated; (B) Alignment of the RNAi transgene regions. Lines between nucleotides indicate identity, identical letters between amino acid sequences indicate identity. The symbol (+) indicates an amino acid substitution by a similar residue, while a (0) or a blank between nucleotide or amino acid letter, respectively, indicates a non-similar substitution. Bottom: amino acid alignment.

Figure 2

Predicted sequence and translation from Coleomegilla maculata transcriptomes most similar to Leptinotarsa decemlineata β-actin used to design beetle resistant potato. (A) Complete cDNA sequence. Highlighted section is the portion of the sequence matching the potato RNAi transgene. Upper case indicates translated sequence, lower case untranslated. Start sites are underscored in unaligned nt sequences; (B) Alignment of the RNAi transgene regions. Translation start sites are boxed. Lines between nucleotides indicate identity, highlighting indicates continuous regions of nucleotide identity. Translated sequence is not included with nucleotides and translation is shown without alignment because all translated residues are identical.

Figure 2

Predicted sequence and translation from Coleomegilla maculata transcriptomes most similar to Leptinotarsa decemlineata β-actin used to design beetle resistant potato. (A) Complete cDNA sequence. Highlighted section is the portion of the sequence matching the potato RNAi transgene. Upper case indicates translated sequence, lower case untranslated. Start sites are underscored in unaligned nt sequences; (B) Alignment of the RNAi transgene regions. Translation start sites are boxed. Lines between nucleotides indicate identity, highlighting indicates continuous regions of nucleotide identity. Translated sequence is not included with nucleotides and translation is shown without alignment because all translated residues are identical.