Development of EST−SSR Markers and Generalizability Analysis of Cinnamomum officinarum Based on Transcriptome Sequence

SSR loci were detected in 187095 unigenes of Cinnamomum officinarum obtained by transcriptome sequencing using MISA software to explore the distribution pattern and compositional characteristics of SSRs and design primers for primer polymorphism and generalization analysis. The results showed that 90590 SSR sites were found in 187095 unigenes, and the frequency of SSR occurrence was 48.42%, with an average of 1 SSR site in 1.99 kb. The total number of unigenes containing SSR sites was 68573, and the frequency of SSR occurrence was 36.65%. Cinnamomum officinarum EST-SSR repeat types had the highest frequency of occurrence of dinucleotide repeat types, followed by trinucleotide, in addition to mononucleotide. Among the 21,777 dinucleotide repeat SSRs detected, repeat unit AG/CT(16.06%) had the highest number and was the dominant repeat unit, followed by AT/AT(5.37%), AC/GT(2.53%), and CG/CG(0.09%). The trinucleotide AAG/CTT (4.37%) had the highest frequency of occurrence and was the dominant repeating unit. Primer3.0 was used to design and synthesize 118 pairs of SSR primers, 96 pairs were amplified successfully(81.36%), of which 14 pairs showed polymorphism(11.86%), and the average polymorphism information content(PIC) was 0.378 in 5 different chemotypes with 3 pairs of primers associated with highly polymorphic sites. The 14 pairs of EST−SSR primers developed in this study showed 100% generalizability in 5 species of the same genus, including C. longepaniculatum, C. kanehirae, C. parthenoxylon, C. burmanni and C. bodinieri. In conclusion, the EST-SSR markers developed based on the transcriptome of C. officinarum have high polymorphism and generalizability, and the method is feasible.