Herbgenomics provides a global platform to explore the genetics and biology of herbs on the genome level. Panax ginseng C.A. Meyer is an important medicinal plant with numerous pharmaceutical effects. Previous reports mainly discussed the transcriptome of ginseng at the organ level. However, based on mass spectrometry imaging analyses, the ginsenosides varied among different tissues. In this work, ginseng root was separated into three tissues—periderm, cortex and stele—each for five duplicates. The chemical analysis and transcriptome analysis were conducted simultaneously. Gene-encoding enzymes involved in ginsenosides biosynthesis and modification were studied based on gene and molecule data. Eight widely-used ginsenosides were distributed unevenly in ginseng roots. A total of 182,881 unigenes were assembled with an N50 contig size of 1374 bp. About 21,000 of these unigenes were positively correlated with the content of ginsenosides. Additionally, we identified 192 transcripts encoding enzymes involved in two triterpenoid biosynthesis pathways and 290 transcripts encoding UDP-glycosyltransferases (UGTs). Of these UGTs, 195 UGTs (67.2%) were more highly expressed in the periderm, and that seven UGTs and one UGT were specifically expressed in the periderm and stele, respectively. This genetic resource will help to improve the interpretation on complex mechanisms of ginsenosides biosynthesis, accumulation, and transportation.