Currently, DNA and RNA are used separately to capture different types of gene mutations. DNA is commonly used for the detection of SNVs, indels and CNVs; RNA is used for analysis of gene fusion and gene expression. To perform both DNA sequencing (DNA-seq) and RNA-seq, material is divided into two copies, and two different proce-dures are required for sequencing. Due to overconsumption of samples and experimental process complexity, it is necessary to create an experimental method capable of analyzing SNVs, indels, fusions and expression. We developed an RNA-based hybridization capture panel targeting actionable driver oncogenes in solid tumors and corresponding sample preparation and bioinformatics workflows. Analytical validation with an RNA standard reference containing 16 known fusion mutations and 6 SNV mutations demonstrated a detection specificity of 100.0% [95% CI 88.7%~100.0%] for SNVs and 100.0% [95% CI 95.4%~100.0%] for fusions. The targeted RNA panel achieved a 0.73-2.63 copies/ng RNA lower limit of detection (LOD) for SNVs and 0.21-6.48 copies/ng RNA for fusions. Gene expression analysis revealed a correlation greater than 0.9 across all 15 cancer-related genes between the RNA-seq re-sults and targeted RNA panel. Among 1253 NSCLC FFPE tumor samples, multiple mutation types were called from DNA- and RNA-seq data and compared between the two assays. The DNA panel detected 103 fusions and 21 METex14 skipping events; 124 fusions and 26 METex14 skipping events were detected by the target RNA panel; 21 fusions and 4 METex14 skipping events were only detected by the target RNA panel. Among the 173 NSCLC samples negative for targetable mutations by DNA-seq, 15 (15/173, 8.67%) showed targetable gene fusions that may change clinical decisions with RNA-seq. In total, 226 tier I and tier II missense variants for NSCLC were analyzed at ge-nomic (DNA-seq) and transcriptomic (RNA-seq) levels. The positive percent agreement (PPA) was 97.8%, and the positive predictive value (PPV) was 98.6%. Interestingly, var-iant allele frequencies were generally higher at the RNA level than at the DNA level, suggesting relatively dominant expression of mutant alleles. PPA was 97.6% and PPV 99.38% for EGFR 19del and 20ins variants. We also explored the relationship of RNA expression with gene copy number and protein expression. The RPKM of EGFR transcripts assessed by the RNA panel showed a linear relationship with copy number quantified by the DNA panel, with an R of 0.8 in 1253 samples. In contrast, MET gene expression is regulated in a more complex manner. In IHC analysis, all 3+ samples exhibited higher RPKM levels; IHC level of 2+ and below showed lower RNA expression. Parallel DNA- and RNA-seq and systematic analysis demonstrated the accuracy and robustness of the RNA sequencing panel in identifying multiple types of variants for cancer therapy.

25 26 Running title: E. corollae genetic structure 27 2 28 Abstract: The hoverfly, Eupeodes corollae, is a worldwide natural enemy of aphids and a 29 plant pollinator. To provide insights into the biology of this species, we examined its 30 population genetic structure by obtaining 1.15-GB random genomic sequences using next-31 generation sequencing and developing genome-wide microsatellite markers. A total of 32 79,138 microsatellite loci were initially isolated from the genomic sequences; after strict 33 selection and further testing of 40 primer pairs in eight individuals, 24 polymorphic 34 microsatellites with high amplification rates were developed. These microsatellites were 35 used to examine the population genetic structure of 96 individuals from four field 36 populations collected across southern to northern China. The number of alleles per locus 37 ranged from 5 to 13 with an average of 8.75; the observed and expected heterozygosity 38 varied from 0.235 to 0.768 and from 0.333 to 0.785, respectively. Population genetic 39 structure analysis showed weak genetic differentiation among the four geographical 40 populations of E. corollae, suggesting a high rate of gene flow reflecting likely widespread 41 migration of E. corollae in China. 42 43 3 46 Introduction 47 Eupeodes corollae is one of the most common hoverflies with a worldwide distribution [1, 2]. 48 The larval stage of this species is mostly insectivorous, feeding mainly on aphids [3-5] while 49 adults are pollinators [6][7][8]. Many hoverfly species are important biological control agents of 50 aphids due to their rapid dispersal and absence of summer diapause compared with other 51 aphidophaga [9]. Understanding the biology and behavior of hoverflies can help in assessing 52 their potential as biological control agents of aphids. 53Hoverflies migrate seasonally as revealed by radar monitoring [10] and isotopic tools 54 [11]. Population genetic analysis is also frequently employed to reveal the migration of 55 species as a complementary approach to traditional methods [12][13][14][15]. In populations of the 56 hoverflies Cheilosia longula [16], Blera fallax [17], Sphaerophoria scripta and Episyrphus 57 balteatus [18], population genetic differentiation has not been found between some regions, 58 suggesting migratory movements of these hoverflies between regions including southern 59 and northern regions of Europe [18, 19]. However, some hoverflies, such as E. balteatus 60 and Scaeva selenitica, are only partially migratory [20]. 61 Previous studies reported that E. corollae is a highly migratory species in Europe [21-62 23], but its migratory behavior of E. corollae remains unclear in other areas. E. corollae is 63 commonly found across China, but the ecology and biology of this species has rarely been 64 studied [8]. In this study, we conducted a preliminary examination of the population genetic 65 structure of E. corollae in China. First, we obtained random genomic sequences of E. 66 corollae using next-generation sequencing and developed an...