The bla CTX-M-15 gene primers aligned with positions 3-876 of accession number FJ815288.
Three criteria were established to filter the comparisons: (i) high degree of similarity (> 90%) between primer and aligned sequence; (ii) primers aligning with only one sequence containing an SSR; (iii) a hypothetical PCR product size longer than 100 bp.
Intron and flanking exon sequences from primer to primer were aligned with Muscle [ 33] and trimmed to remove ambiguous sites.
12, 13 The PLU5/6 primers were aligned with the 18S sequence of Z. lanceolata.
Using the re-sequencing reads from 1765 rice varieties as the template, unique primer sequences were aligned with Bowtie, allowing up to one mismatch (Fig. 6 Step 3).
The obtained sequences, except for the primer regions, were aligned with the corresponding sequences of other rickettsiae deposited in the GenBank database to identify known sequences with a high degree of similarity using multisequence alignment programs, the Phydit software (10 ), and the MegAlign software package (Windows version 3.12e; DNASTAR, DYNASTAR Inc., Madison, WI, USA).
Forward cross primer (sense) and reverse cross primer (anti-sense) are aligned with the 5′ end of the sequence.
Elimination of genomic DNA from cDNA preparation was verified by PCR with primers aligned in different exons for gene Translation elongation factor (GmELF1A) and Glucose-6-phosphate dehydrogenase (GmG6PD) as described by Miranda et al. [[ 41]].
Sequences amplified from the same primer set were initially aligned with CLC Sequence Viewer 6.9.1 (CLC Bio, Valencia, CA, USA) for preliminary SNP verification.
Sequences were obtained using the ABI 310 genetic analyser (Applied Biosystems, Bedford, MA, USA) with proper primers, and they were aligned with known CMV variants in GenBank afterward.
