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Background  Polyploidy is very common in plants and can be seen as one of the key drivers in the domestication of crops and the establishment of important agronomic traits. It can be the main source of genomic repatterning and introduces gene duplications, affecting gene expression and alternative splicing. Since fully sequenced genomes are not yet available for many plant species including crops, de novo transcriptome assembly is the basis to understand molecular and functional mechanisms. However, in complex polyploid plants, de novo transcriptome assembly is challenging, leading to increased rates of fused or redundant transcripts. Since assemblers were developed mainly for diploid organisms, they may not well suited for polyploids. Also, comparative evaluations of these tools on higher polyploid plants are extremely rare. Thus, our aim was to fill this gap and to provide a basic guideline for choosing the optimal de novo assembly strategy focusing on autotetraploids, as the scientific interest in this type of polyploidy is steadily increasing.    Results  We present a comparison of two common (SOAPdenovo-Trans, Trinity) and one recently published transcriptome assembler (TransLiG) on diploid and autotetraploid species of the genera  Acer  and  Vaccinium  using  Arabidopsis thaliana  as a reference. The number of assembled transcripts was up to 11 and 14 times higher with an increased number of short transcripts for  Acer  and  Vaccinium , respectively, compared to  A. thaliana.  In diploid samples, Trinity and TransLiG performed similarly good while in autotetraploids, TransLiG assembled most complete transcriptomes with an average of 1916 assembled BUSCOs vs. 1705 BUSCOs for Trinity. Of all three assemblers, SOAPdenovo-Trans performed worst (1133 complete BUSCOs).    Conclusion  All three assembly tools produced complete assemblies when dealing with the model organism  A. thaliana , independently of its ploidy level, but their performances differed extremely when it comes to non-model autotetraploids, where specifically TransLiG and Trinity produced a high number of redundant transcripts. The recently published assembler TransLiG has not been tested yet on any plant organism but showed highest completeness and full-length transcriptomes, especially in autotetraploids. Including such species during the development and testing of new assembly tools is highly appreciated and recommended as many important crops are polyploid.    Supplementary Information  The online version contains supplementary material available at 10.1186/s12859-021-04078-8.

Comparing de novo transcriptome assembly tools in di- and autotetraploid non-model plant species