Diversity analysis of reverse transcriptase gene (RT) of long terminal repeat retrotransposons in Arachis ipaensis Krapov. et W. C. Greg. with BB genome

The reverse transcriptase gene (RT) sequences of Ty1-copia-like (Type 1) and Ty3-gypsy-like retrotransposons (Type 2) were amplified and isolated from the Arachis ipaensis Krapov. et W. C. Greg. with BB genome by degenerate polymerase chain reaction (PCR). Sequence characteristics, diversity, phylogenetic relationships, and transcriptional activity were then analyzed. Results showed that the target bands for types 1 and 2 were 260 bp and 430 bp in size, respectively. 23 sequences of type 1 retrotransposons and 32 sequences of type 2 retrotransposons were obtained. Sequence lengths of type 1 and type 2 ranged from 262 to 266 bp and from 395 to 435 bp, respectively. The proportion of AT content in type 1 and type 2 ranged from 61.60% to 69.17% and from 55.79% to 61.34%, respectively. Similarity between nucleotide sequences in type 1 and type 2 ranged from 52.5% to 98.9% and from 45.0% to 98.8%, respectively. The similarity between amino acid sequences of type 1 and type 2 ranged from 39.8% to 100% and from 9.0% to 97.2%, respectively. The heterogeneity of type 2 was higher than that of type 1. There were three and 15 nonsense mutations in type 1 and type 2, respectively. The incidence of nonsense mutations in type 2 was much higher than that in type 1. The conserved motifs of type 1 were highly conserved, while the conserved motifs of type 2 showed a certain degree of variation. The protein tertiary structures were similar in overall configuration. Type 2 had more differences in protein structure than type 1. Based on cluster analysis, type 1 and type 2 were divided into five and six families, respectively. The phylogenetic tree showed that type 1 and type 2 were divided into four and 11 classes, respectively, but type 2 sequence categories and diversity were significantly higher than that of type 1. Several type 1 and 2 RT gene sequences were closely related to the RT gene sequences of other species, indicating possible horizontal transmission of retrotransposons. When searching the peanut EST database, one type 1 and seven type 2 sequences from the A.ipaensis BB genome showed transcriptional activity. The type 2 retrotransposons showed greater transcriptional activity than the type 1 retrotransposons. This study not only provides sequences for the isolation of full-length LTR retrotransposons and for studies on their transcriptional activity and function, but also lays the foundation for the development of molecular markers based on LTR retrotransposons in the Arachis genus.