Missing (MP) and functionally uncharacterized proteins (uPE1) comprise less than 5% of the total number of proteins encoded by human Chr18 genes. Within half a year, since the January 2020 version of NextProt, the number of entries in the MP+uPE1 datasets changed, mainly due to the achievements of antibody-based proteomics. Assuming that the proteome is closely related to the transcriptome scaffold, quantitative PCR, Illumina HiSeq, and Oxford Nanopore Technology were applied to characterize the liver samples of three male donors in comparison with the HepG2 cell line. The data mining of the Expression Atlas (EMBL-EBI) and the profiling of biopsy samples by using orthogonal methods of transcriptome analysis have shown that in HepG2 cells and the liver, the genes encoding functionally uncharacterized proteins (uPE1) are expressed as low as for the missing proteins (less than 1 copy per cell), except the selected cases of HSBP1L1, TMEM241, C18orf21, and KLHL14. The initial expectation that uPE1 genes might be expressed at higher levels than MP genes, was compromised by severe discrepancies in our semi-quantitative gene expression data and in public databanks. Such discrepancy forced us to revisit the transcriptome of Chr18, the target of the Russian C-HPP Consortium. Tanglegram of highly expressed genes and further correlation analysis have shown the severe dependencies on the mRNA extraction method and the analytical platform. Targeted gene expression analysis by quantitative PCR (qPCR) and high-throughput transcriptome profiling (Illumina HiSeq and ONT MinION) for the same set of samples from normal liver tissue and HepG2 cells revealed the detectable expression of 250+ (92%) protein-coding genes of Chr18 (at least one method). The expression of slightly more than 50% protein-coding genes was detected simultaneously by all three methods. Correlation analysis of the gene expression profiles showed that the grouping of the datasets depended almost equally on both the type of biological material and the experimental method, particularly cDNA/mRNA isolation and library preparation.