[Expression of Leafy Cotyledon 2 from Arabidopsis increased the content of lipid in Chlorella sorokiniana].

The low lipid content is one of the major bottlenecks to realize the industrialization of the algae biodiesel. Improvement of lipid content through global regulation to get high-yield generating algae is a good strategy. Leafy Cotyledon 2 (LEC2) is an important transcription factor for seed maturation and oil accumulation in Arabidopsis. However, there are few reports regarding adoption of LEC2 for lipid accumulation until now. In this study, LEC2 from Arabidopsis was cloned into the plant expression vector pCIMBIA1300 and transformed into C. sorokiniana through particle bombardment. One recombinant was screened by PCR, RT-PCR and Western blot analyses. Compared with the wild type one, the total lipid content in the recombinant increased one fold, which did not show effect on cell growth, indicating that LEC2 can efficiently enhance the lipid accumulation in C. sorokiniana.

Flanking sequence determination and event specific detection of transgenic wheat B72-8-11b strain.

Exogenous fragment sequence and flanking sequence between exogenous fragment and recombinant chromosome of transgenic wheat B72-8-11b were successfully acquired through PCR amplification with cross-matched primers from exogenous genes. Newly acquired exogenous fragment covered the full-length sequence of transformed genes such as transformed plasmid and corresponding functional genes including marker uidA, promoter ubiquitin, lacZ, 1Dx5, and part of sequence of the wheat genome. A specific PCR detection method for transgenic wheat B72-8-11b strain was established on the basis of primers designed according to flanking sequence. The designed primers revealed specific amplification of 132 bp product of transgenic wheat B72-8-11b strain. This method is characteristics of high specificity, high reproducibility, rapid identification, and excellent accuracy for the identification of transgenic wheat B72-8-11b strain.

Differential fucosyltransferase IV expression in squamous carcinoma cells is regulated by promoter methylation.

Enhanced fucosyltransferase IV (FUT4) expression correlates with increased tumor malignancy in many carcinomas. However, little is known about the regulation of FUT4 expression, and whether FUT4 expression is influenced by the methylation status of the FUT4 promoter is unclear. In this study, we demonstrated that FUT4 expression is negatively correlated with the methylation degree of a CpG island in the FUT4 promoter, suggesting that the methylation status of FUT4 promoter regulates the expression of FUT4. The results indicate that manipulating the methylation status of the FUT4 promoter to regulate FUT4 expression may be a novel approach in the treatment of malignant tumors.

Integrity of nonviral fragments in recombinant Tomato bushy stunt virus and defective interfering RNA is influenced by silencing and the type of inserts.

Recombinant plant viruses have the propensity to remove foreign inserts during replication. This process is virus-specific and occurs in a host-dependent manner. In the present study, we investigated the integrity of foreign inserts in recombinant plant viruses using a model system consisting of Tomato bushy stunt virus (TBSV) and its defective interfering RNA (DI). These were tested in Nicotiana benthamiana plants that were either wild type or transgenic for the green fluorescent protein (GFP) gene. GFP-derived inserts were retained in the recombinant TBSV and DI population that were inoculated onto GFP-transgenic N. benthamiana plants in which silencing of the GFP transgene was initiated, but they were removed from the virus and DIs that were maintained on wild-type plants. A foreign insert derived from an endogenous N. benthamiana gene encoding the H subunit of the magnesium chelatase (NbChlH) was deleted, whereas the fragment of an RNA-dependent RNA polymerase gene (NbRdRP1m) was retained in the recombinant TBSV population. These results demonstrate that the recombination of TBSV to remove nonviral fragments is influenced by silencing and the type of inserts.

A novel co-delivery system consisting of a Tomato bushy stunt virus and a defective interfering RNA for studying gene silencing.

Virus induced gene silencing (VIGS) and suppression are RNA-specific defense and counter-defense circuits in plant-virus interactions. These phenomena have been investigated extensively with an Agrobacterium-mediated transient expression system. In this study, a virus-based transient expression system was developed to study these phenomena. A Tomato bushy stunt virus (TBSV) viral vector with an inactivated P19 suppressor gene, referred to as pHST2-14, was chosen to express the P1 of Tobacco etch virus (TEV). TEV P1 is a component of a well-characterized VIGS suppressor, TEV P1/HC-Pro protein. A TBSV defective interfering RNA (DI) that contains the 3' proximal portion of a green fluorescence protein (GFP) gene, DI-P, was used as a silencing inducer of the homologous GFP gene on GFP transgenic Nicotiana benthamiana (NbGFP) plants. The TEV P1 gene was inserted into pHST2-14 to generate TBSV-P1. Transcripts of TBSV-P1 were then mixed with DI-P transcripts and inoculated onto NbGFP plants. DI-P consistently accumulated in NbGFP plants that were inoculated with TBSV-P1 and DI-P, and efficiently induced silencing of GFP transgene. These results demonstrate that a TBSV-based co-delivery system can provide a new alternative tool to investigate gene silencing and its influence by a TBSV-expressed foreign protein. It also can be used to elucidate functions of endogenous genes in plants.