Cloning and characterisation of the rad9 DNA repair gene from Schizosaccharomyces pombe

The rad9.192 DNA repair mutant from the fission yeast, Schizosaccharomyces pombe, is sensitive to both UV and ionising radiation. The rad9 gene has been cloned by complementation of the gamma-ray sensitivity of the mutant cell line. A 4.3kb HindIII fragment was found to confer resistance to both types of radiation. The region of complementation was further localised to a 2.6kb HindIII-EcoRV fragment, which, by DNA sequence analysis, was found to contain sequences capable of coding for a 427 amino acid protein, if three introns were postulated to remove stop codons. The introns were confirmed by sequence analysis of cDNA clones and PCR products derived from cDNA. The product of transcription is a 1.6kb mRNA of low abundance. The putative rad9 protein shows no homology to any published sequence. A truncated protein is capable of complementing the radiation sensitivity of the rad9.192 mutant. Deletion of the gene is not lethal and the null allele has a similar phenotype to the rad9.192 mutant

Evolutionary conservation of excision repair in Schizosaccharomyces pombe: Evidence for a family of sequences related to the Saccharomyces cerevisiae RAD2 gene

Cells mutated at the rad13 locus in the fission yeast, Schizosaccharomyces pombe are deficient in excision-repair of UV damage. We have cloned the S.pombe rad13 gene by its ability to complement the UV sensitivity of a rad13 mutant. The gene is not essential for cell proliferation. Sequence analysis of the cloned gene revealed an open reading-frame of 1113 amino acids with structural homology to the RAD2 gene of the distantly related Saccharomyces cerevisiae. The sequence similarity is confined to three domains, two close to the N-terminus of the encoded protein, the third being close to the C-terminus. The central region of about 500 amino acids shows little similarity between the two organisms. The first and third domains are also found in a related yet distinct pair of homologous S.pombe/S.cerevisiae DNA repair genes (rad2/YKL510), which have only a very short region between these two conserved domains. Using the polymerase chain reaction with degenerate primers, we have isolated fragments from a gene homologous to rad13/RAD2 from Aspergillus nidulans. These findings define new functional domains involved in excision-repair, as well as identifying a conserved family of genes related to RAD2

Cloning and characterisation of the S.pombe rad15 gene, a homologue to the S.cerevisiae RAD3 and human ERCC2 genes

The RAD3 gene of Saccharomyces cerevisiae encodes an ATP-dependent 5' - 3' DNA helicase, which is involved in excision repair of ultraviolet radiation damage. By hybridisation of a Schizosaccharomyces pombe genomic library with a RAD3 gene probe we have isolated the S.pombe homologue of RAD3. We have also cloned the rad15 gene of S.pombe by complementation of radiation-sensitive phenotype of the rad15 mutant. Comparison of the restriction map and DNA sequence, shows that the S.pombe rad15 gene is identical to the gene homologous to S.cerevisiae RAD3, identified by hybridisation. The S.pombe rad15.P mutant is highly sensitive to UV radiation, but only slightly sensitive to ionising radiation, as expected for a mutant defective in excision repair. DNA sequence analysis of the rad15 gene indicates an open reading frame of 772 amino acids, and this is consistent with a transcript size of 2.6kb as detected by Northern analysis. The predicted rad15 protein has 65% identity to RAD3 and 55% identity to the human homologue ERCC2. This homology is particularly striking in the regions identified as being conserved in a group of DNA helicases. Gene deletion experiments indicate that, like the S.cerevisiae RAD3 gene, the S.pombe rad15 gene is essential for viability, suggesting that the protein product has a role in cell proliferation and not solely in DNA repair

Orbital angular momentum exchange in an optical parametric oscillator

We present a study of orbital angular momentum transfer from pump to down-converted beams in a type-II Optical Parametric Oscillator. Cavity and anisotropy effects are investigated and demostrated to play a central role in the transverse mode dynamics. While the idler beam can oscillate in a Laguerre-Gauss mode, the crystal birefringence induces an astigmatic effect in the signal beam that prevents the resonance of such mode.Comment: 10 pages, 8 figures, regular articl

The effect of 2,4-D, BA and Thidiazuron on somatic embryo induction of liberica coffee of Tungkal Composite from Jambi

The Tungkal Composite of liberica coffee rejuvenation program at Jambi Province is urgently needed due to the latest status of the plants are old and showing diseased attacked. To support the rejuvenation, the provision of seedlings could be done by tissue culture approach through somatic embryogenesis. This research aimed to study the effect of 2,4-D, BA and TDZ on callus formation and embryo induction from liberica coffee leaf explants. Explants used were leaf sections with or without veins. Explants were grown on Murashige Skoog (MS) basal medium with 2,4-D + BA and 2,4-D + TDZ. A completely randomized design was employed in the trial, and each experimental unit was replicated three times. Results showed that callus could be formed on both types of explants, but leaf explants with veins could produce callus faster. The use of BA in medium could induce callus formation faster than TDZ. The higher the concentration of 2,4-D, BA and TDZ, the slower callus initiation. The addition of TDZ in the medium resulted in mostly callus, while callus from medium with BA showed a friable structure. Globular somatic embryos were formed in all treatments with varying amounts. TDZ can induce more globular somatic embryo than BA