Xanthopappus subacaulis C. Winkl., endemic to the Qinghai-Xizang Plateau, is a perennial medicinal plant from the genus Xanthopappus of the family Asteraceae, with the important economic, ecological and medicinal values. In order to determine the appropriate sequencing strategy for the whole genome of X. subacaulis, we analyzed and evaluated the genome size, heterozygosity, repeatability and GC content based on the flow cytometry and survey analysis method in the present study. The result was as follows: (1) Using Opisthopappus longilobus and Solanum lycopersicum as reference species, flow cytometry was used to estimate that X. subacaulis is a diploid, with genome sizes of 1.94 G or 1.75 G, and a DNA-C value of 0.99 pg. The genome size (G) of approximately was 0.99 pg, which was estimated through flow cytometry. (2) The high-throughput sequencing generated approximately 106.6 GB of raw data. After quality control, the Q20 and Q30 of the clean reads were greater than 97.1% and 90.8%, respectively. Besides, there was no significant separation of base contents within AT and GC, and the proportion of GC content was 38.5%. It indicated that the second-generation sequencing quality was good. (3) Based on K-mer analysis, the genome size of X. subacaulis was 2198.50 Mb, the heterozygosity was 0.69%, and the proportion of repeat sequences was 80.15%, which suggested that it belonged to the complex genome with micro-heterozygosity and high repeat sequence. (4) The identification results of LTR-REs indicate that the Copia family is the most abundant, accounting for 30.72% of the whole genome, while the Gypsy family and Unknown account for 33.66% and 16.54%, respectively. Moreover, their insertion time began approximately 3 Mya, with a significant amplification occurring within the last 1 Mya. Therefore, this study suggests that the large-scale insertion of LTR elements is one of the important factors leading to the genomic complexity of X. subacaulis. Thus, we thought that the de novo whole-genome sequencing of X. subacaulis should be adopted the sequencing and assembly strategy with the mixture of the second and third generation sequencing. This study provided the reference data for the genetic map construction of high-quality and key functional gene-mining for X. subacaulis.