Genome variation in colorectal cancer patient with liver metastasis measured by whole-exome sequencing.

BACKGROUND: Liver metastasis of colorectal cancer (CRC) is an important cause of death from CRC, but its molecular mechanism is still unclear. In recent years, whole-exome sequencing has played an increasingly important role in the study of the occurrence and development of diseases, especially malignant tumors. Its high throughput and low cost advantages enable researchers to explore the pathogenic genes of diseases, and screen potential molecular markers and therapeutic targets from the level of genomics. METHODS: This study collected the primary tumor tissues, matched paracancerous, normal tissues, and liver metastases of 4 CRC patients admitted to the Department of General Surgery of the First Affiliated Hospital of Soochow University, and performed high-depth whole-exome sequencing, with the sequencing depth of each sample reaching 123× on average, then filtered the sequencing data, compared them, and analyzed the bioinformatics data. RESULTS: we found 8,565 single nucleotide variants (SNV) and 429 insertions/deletions (InDel) in the primary and hepatic lesion tissues, and the genes with the highest mutation frequency were titin (TTN), obscurin (OBSCN), and homeodomain-interacting protein kinase 2 (HIPK2). The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the mutant genes was conducted, and it was found that the mutant genes were mainly concentrated in the cells, cell parts, and cellular process of GO. The results of KEGG pathway analysis showed that mutations were mainly distributed in circadian entrainment, insulin secretion, and glutamatergic synapse. Further, we identified 723 SNV and Indel genes with high frequency mutations including TTN, OBSCN, and hydrocephalus-inducing protein homolog (HYDIN) across all tissues of liver metastases. The GO analysis showed that the mutated genes in liver metastatic tissues were mainly concentrated in cell, cell part, and cellular process. The KEGG pathway analysis showed that high frequency mutation genes were focused on gastric acid secretion, bile secretion, and melanogenesis. CONCLUSIONS: This study found some candidate genes related to the occurrence of CRC and liver metastasis through whole-exome sequencing of relevant tissues in CRC patients with liver metastasis, which is expected to provide new markers and therapeutic targets for such patients.

[Raising fat content in transgenic rice by anti-PEP gene transformation].

Gene for phosphoenolpyruvate carboxylase (PEPCase) was incorporated in antisense orientation into genome of japonica rice variety Xiushui 11 through Agrobacterium tumefaciens-mediated transformation of mature seeds-derived embryogenic calli, and a total of 95 transgenic plants were regenerated, which were confirmed by histochemical detection of GUS activity (Fig.2), PCR assay (Fig.3) and Southern hybridization analysis (Fig.4). The transgene was inherited by the T1 progeny and Mendelian 3:1 segregation ratio was observed in most transgenic lines. It was found that the fat content of hulled grains of T2 transgenic lines was 0.37 +/- 0.12 per cent higher than that of the control, most of which were statistically very significant (Table 2).

Highly efficient transformation and plant regeneration of tall fescue mediated by Agrobacterium tumefaciens.

An efficient and reproducible system has been developed for the production of transgenic plants in tall fescue (Festuca arundinacea Schreb.) using A. tumefaciens-mediated transformation. Two-months-old suspension cultures served as excellent explants for transformation. The explants were inoculated with an A. tumefaciens strain EHA105 carrying a plasmid pDBA121 containing genes for hygromycin phosphotransferase (hpt) and phosphinotricin acetyltransferase (bar). The commercial herbicide Basta was used as a selective agent. Inclusion of acetosyringone (ACS) 20 mg/L in the co-culture medium led to an increase in transformation efficiency. The efficiency of transformation was highly dependent on the genotype, the explant, the culture medium and selective agent used. Tall fescue transformation efficiency is 2.85-10.9 plants per gram fresh weight (FW) of suspension cultures. This is much higher than the corresponding values reported before (2-5 plants). So far more than 300 transgenic plants have been obtained, the fertility of some transgenic plants had decreased. Stable integration and high expression of the transgenes were confirmed by PCR analysis and Southern blot hybridization or herbicide Basta spraying test.