Transcription factors and developmental control in plants

Lots of plant transcription factors have been identified in the recent decade, notably the knotted-1 related transcription factor family, the MADS-box transcription factors family, and factors involved in induction of floral meristem and those controlling cellular differentiation in plants. These transcription factors not only play an important role in the growth, development,

Differential display of carrot radicle genes in different developmental states

mRNA differential display was used in analyzing the expression of carrot somatic embryo radicle gene in different developmental stages brought about by controlled cultivation. As shown by experiment, marked differences in electrophoresis patterns of DNA exist between the normally developing carrot radicle and those with their development inhibited. 12 series of specific bands have been cloned, and structural analysis of them is under way.

Isolation of tomato proteinase inhibitor Ⅱ gene and the function of its intron

The genomic DNA sequence of tomato pro-teinase inhibitor II gene (named tin2i, whose accession number in GenBank is AF007240) was isolated by PCR techniques. The intron sequence (TPI), with a length of 109 bp, owns typical structures of GT/AG dinucieotides at both ends and high content of AT base pairs which accounts for 80.7% of the total nucleotides. As shown by recombination experiment, the TPI sequence could efficiently promote the expression of the reporter gene gusA and this effect was independent of the position and orientation of the intron, thus showing its role as an enhancer. Such experiments as gel retardation assays, GUS histochemical staining and GUS fluorometric assays further demonstrated that TPI sequence maybe has promoter-like activity.

Application of a nuclear localization signal gene in transgene mice

Efficient gene transfer by cytoplasm co-injec-tion will offer a powerful means for transgenic animals. Us-ing co-injection in cytoplasm, two independent gene con-structs, including bovine a-s1-casein-hG-CSF and a mammal expression vector expressing a nuclear localization signal (mNLS), were introduced into fertilized mouse eggs. The target gene construct was docked into host nucleus probably by the nuclear localization signal. Transgene mice have been obtained at 58% (29/50) of integration ratio. Ex-pression level of the positive transgene mice was detected by Western blotting. Maximal expression of human G-CSF was estimated about 540 mg/L of milk. The expression ratio was up to 75% (9/12). The results here have important practical implications for the generation of mammary gland bioreac-tors and other transgene studies. Co-injection of a target gene with an expression vector of a mammal nuclear localization signal by cytoplasm appears to be a useful, efficient and easy strategy for generating

Mutational effect of the “-35” element of sorghum psbA gene promoter

Mutations of the first position T and the third position G in TTGACA, the ' - 35' element of sorghum psbA gene promoter, were induced using chemically synthesized 20 nt oligonucleotide primer. Three mutants were produced: ATTACA, GTGACA, and ATGACA. Then the protein binding affinity of the mutants and the wild type sorghum psbA gene promoter was tested in a spinach chloroplast protein extract system. Gel retardation assay of the

Establishment and application of both FLP and Cre site-specific recombination systems at the same position in the genome

Both FRT-FRT and LoxP-LoxP sites that are the target sepuences of site-specific recombinases have been constructed in a vector, called C4LFY, using the recombinant DNA technigue. C4LFY also contains P elements, 2 exons and 1 intron of Drosophila yellow gene, yellow promoter and enhancers, and flanking DNA. Since C4LFY made use of two pairs of FRT and LoxP sites, this vector included two site-specific recombination systems. C4LFY was then integrated into Drosophila genome by P-element-mediated germ line transformation. in the presence of the FLP or Cre recombinase, either FLP/FRT or Cre/LoxP recombination reaction was successfully created at the same position in the genome. Using this system, the molecular basis of yellow gene expression and regulation during development have been investigated. Results indicate that the tissue-specific expression of yellow gene is directly regulated by transcriptional enhancers. in addition, the 5’ and 3’ genomic sequences flanking the yellow gene have been

Expression of self-incompatibility ribonucleases of Antirrhinum in Escherichia coli

Self-incompatibility is an intraspecific reproductive barrier to prevent self-fertilization inthe flowering plants. In many species, self-incompatibility is controlled by a single S locus with multiple alleles. So far, the only gene known in the S locus of the Solanaceae, Scrophulariaceae and Rosaceae encodes a class of ribonucleases, called self-incompatibility ribonucleases (S RNases), which have been shown to mediate stylar expression of self-incompatible reaction. As the first step to investigate their three-dimensional structure, we successfully expressed three biologically active S RNases of Antirrihnum (S2, S4 and S5) in Escherichia coli (E. coli). Their functional expressions caused no detrimental effect on host bacteria growth and provided a basis for a large scale preparation of S RNase proteins. Possible reasons for non-lethality of S RNases on E. coliare discussed.

Isolation of candidate R disease resistance genes from rice

Using a polymerase chain reaction (PCR) based method six distinct candidate disease resistant gene (R) homologs from rice have been isolated. The rice sequences are organized into two phylogenetic groups with contrasting genomic organization patterns. The first group, represented by a single sequence, Osh359-1, is more similar to non-rice R sequences than to rice ones and has a simple genomic organization. The second group, represented by Osh359-3, contains the remaining five rice sequences. Osh359-3 consists of a multi-gene family. The members of Osh359-3 family are further found to be clustered together in the genome.

Isolation of cDNA fragment of fertility-related gene inphotoperiod-sensitive genic male sterile rice

The photoperiod_sensitive genic male sterile rice (PGMR) is particularly useful to take advantage of heterosis in rice. mRNA differential display was used to isolate the fertility_relative genes in rice. After establishing an optimized mRNA differential display system, one of the differential cDNA fragments that maybe related to the development and maturation of rice panicle was cloned from a PGMR Nongken 58S.

Novel gene expressed during early embryogenesis of zebrafish identified by mRNA differential display

Following the revelation of the molecular mechanism of morphogenesis in fruitfly, research on the molecular mechanism of morphogenesis in vertebrate becomes the focus of developmental biology. The isolation of genes controlling the embryogenesis of zebrafish, a vertebrate model animal, is considered as an initial step toward investigating this issue. There are several approaches that can be used to isolate developmental genes, each of which is suited to a particular situation. In this note, mRNA differential display was utilized to demonstrate the mRNA differences among zebrafish embryos at 4, 5 and 6 h post fertilization (28.5℃, corresponding to oblong, dome and shield stages, respectively, called blastula, gastrula and neurula in this note). One cDNA tag that was specific to embryos at neurula stage was cloned and sequenced. After sequence comparison in Genbank, we found that this cDNA tag represents a novel gene. The expression of this gene in the developing zebrafish embryos was examined by whole mount in situ hybridization. The hybridization results confirmed that this gene was specifically expressed in zebrafish neurula embryos.