Abstract: The transcriptome of Asplenium nidus was sequenced by Illumina HiSeq 2000 platform that is a new generation of high-throughput sequencing technology to study the expression profiling and predict the functional genes. The target sample sequencing，a total of 29 254 595 reads fragment contains 5 908 586 517 bp in sequence information were generated. A total of 42 907 unigenes contains 40.16 Mb in sequence information were formed by initial sequence splicing，with an average read length of 936 bp. 24 993 unigenes were annotated using BLASTX searches against the Nr and SwissProt databases. In this study，all assembled unigenes can be broadly divided into biological processes，cellular components and molecular function categories of 51 branches by gene ontology，including metabolic process，binding，catalytic activity and cellular process. Unigenes were further annotated based on COG category，which could be grouped into 24 functional categories. KEGG pathway analysis showed that unigenes can be broadly divided into 116 classes according to the function. There were 6 067 SSR in 42 907 unigenes werefound. The types of SSR were analyzed that AG/CT was the highest repeat，following by AC/GT，A/T and AGG/CCT. Based on flank sequence of detected SSR，20 primer pairs were designed and tested for the amplification efficiency and polymorphism. The results showed that 7 primer pairs showed polymorphism among different fern varieties.

Key words: Asplenium nidus, transcriptome, bioinformatics, gene annotation, simple sequence repeat