Archaea is now recognized as the third domain of life. Since their discovery, much effort has been directed towards understanding the molecular biology and biochemistry of Archaea. The objective is to comprehend the complete structure and the depth of the phylogenetic tree of life. DNA replication is one of the most important events in living organisms and DNA polymerase is the key enzyme in the molecular machinery which drives the process. All archaeal DNA polymerases were thought to belong to family B. This was because all of the products of pol genes that had been cloned showed amino acid sequence similarities to those of this family, which includes three eukaryal DNA replicases and Escherichia coli DNA polymerase II. Recently, we found a new heterodimeric DNA polymerase from the hyperthermophilic archaeon, Pyrococcus furiosus. The genes coding for the subunits of this DNA polymerase are conserved in the euryarchaeotes whose genomes have been completely sequenced. The biochemical characteristics of the novel DNA polymerase family suggest that its members play an important role in DNA replication within euryarchaeal cells. We review here our current knowledge on DNA polymerases in Archaea with emphasis on the novel DNA polymerase discovered in Euryarchaeota.
Distorted segregation has been repeatedly observed in molecular marker linkage mapping of various plants and animals where distant crosses were made. It may be caused by a semi-lethal factor acting in the filial generations. Although general methods have been developed to find semi-lethal factors, so far no method is available to estimate semi-lethal factors in the organisms, for example Bombyx mori and Drosophila melanogaster, in which no crossing-over occurs in meiosis in either of the sexes. In this report, a method is presented for estimating the recombination values between a semi-lethal factor locus and neighboring dominant molecular markers, e.g. RAPDs and AFLPs, and the relative viability of gametes or zygotes affected by the semi-lethal factor in an F2 population for such species by using the maximum likelihood method associated with the expectation maximization (EM) algorithm. Five selection models of gamete or zygote were considered, and the most likely one was determined by goodness of fit of the observed counts of phenotypes to the expected ones under the models. The method was applied to segregation data of RAPD markers of an F2 population with 101 individuals developed from the `C108' × `p50' cross in silkworm, B. mori. The presence of two semi-lethal factor loci (L1 and L2) located on the linkage groups 22a and 27b causing partial selection was suggested. The semi-lethal factors L1 and L2 were theoretically predicted to affect viability of male gametes and zygotes, respectively.
We discovered a novel retrotransposable element, designated Yokozuna, on the W chromosome of Bombyx mori. The size of this element is 4738 bp, including a 208-bp long terminal repeat (LTR) on one side and a 183-bp LTR on the other. This retrotransposable element is flanked by a 5-bp target site duplication, TAATT. Yokozuna contains a single long open reading frame (ORF) and the whole deduced amino acid sequence of ORF reveals strong homology with copia of Drosophila. Moreover, an alignment analysis of the reverse transcriptase (RT) sequences suggested that the Yokozuna element is the first Bombyx retrotransposable element belonging to the Ty1-copia group. The number of Yokozuna per haploid genome is approximately four copies. Although Yokozuna was discovered on the W chromosome, it is not specific for the W chromosome.
In the silkworm, Bombyx mori, a non-long terminal repeat (non-LTR) retrotransposon, BMC1, is considered to be a LINE (long interspersed nuclear element)-like element. So far, a BMC1 containing two intact open reading frames (ORFs) has not been found. However, we discovered a complete full-length BMC1 on the W chromosome. This BMC1 is 5091 bp and contains a 5' untranslated region (5'-UTR), two intact ORFs, and 3'-UTR which terminates in a poly(A) tail. ORF1 encodes a putative nucleic acid-binding protein, while ORF2 encodes a protein containing an endonuclease domain and a reverse transcriptase domain.
A rice gene bph2 for resistance against brown planthopper (BPH), Nilaparvata lugens Stål, was reported to be recessive and either allelic or closely linked to a dominant BPH resistance gene, Bph1. bph2 was introgressed from an indica resistance donor variety, `IR1154-243', into a japonica breeding line, `Norin-PL4'. A segregation analysis of BPH resistance in F2 and F3 progenies from a cross of a japonica susceptible variety, `Tsukushibare', and `Norin-PL4', however, showed that the resistance gene in `Norin-PL4' behaved as a dominant gene. Genotyping of `Norin-PL4' using 99 RFLP markers covering all 12 rice chromosomes showed that `Norin-PL4' possessed a large segment of chromosome 12 introgressed from `IR1154-243'. Six RFLP markers on the introgressed segment was cosegregated with BPH resistance and bph2 was mapped at 3.5 cM from the closest RFLP marker, G2140. The position of bph2 on the standard `Nipponbare'/`Kasalath' map was at a considerable distance (about 30 cM) from that of Bph1 previously mapped using a different population. Despite this, no susceptible recombinants were obtained in a large number of F3 progeny from crosses between two Bph1 carrier lines and `Norin-PL4'. Problems of dominance/recessiveness and no recombinations between the two loci were discussed.
We isolated and characterized yeast mutants whose growth is sensitive to a local anestheticum tetracaine and, at the same time, temperature sensitive. These mutants were collectively called las mutants (local anestheticum sensitive). The las21 mutants were analyzed in this study. The wild type LAS21 gene was cloned by exploiting temperature sensitivity of the las21 mutants and we found that LAS21 encodes ORF YJL062w which has not been analyzed before. Las21p is putative membrane protein belonging to the major facilitator super family containing plural membrane spanning domains. Complete elimination of the LAS21 ORF did not kill the cells but made their growth temperature sensitive. Interestingly, the complete loss of the LAS21 gene canceled the sensitivity to tetracaine. The ability of the las21 mutants to grow at a higher temperature was recovered in the various media containing an osmotic stabilizer or salts. Furthermore, temperature sensitivity of the las21 mutants was partially suppressed by introduction of PKC1, encoding protein kinase C, on a high copy vector. We found some genetic interactions between LAS21 and Ras/cAMP cascade genes. These results suggest that LAS21 defines unkonwn pathway regulating the stress response of yeast.
Powdery mildew-resistant wheat lines, Tra.R9002 with rye chromatin and Tra.V149 with Haynaldia villosa chromatin, were crossed to combine their different powdery mildew-resistant genes in the wheat genetic background. In the F5 generation of Tra.R9002 × Tra.V149, a highly powdery mildew-resistant line W159 was selected according to its chromosome number 2n = 41 ~ 42. In the F6 line W159-9 which was immune to powdery mildew, one pair of rye chromosomes and one pair of wheat-H. villosa translocated chromosomes were identified simultaneously by multicolor fluorescence genomic in situ hybridization in the wheat genetic background (chromosome number 2n = 42). Through C-banding and a sequential C-banding analysis after genomic in situ hybridization, the rye chromosomes were identified as 5R and the wheat-H. villosa translocated chromosomes were identified as 6VS-6AL. This indicates that W159-9 carries two powdery mildew-resistant genes of Pm4 on 5R and Pm21 on 6VS. The wheat chromosomes absent in W159-9 were identified as the 6AS and 5D. On the other hand, seven out of 17 progenies of another F6 line W159-3 possessed 5R and 6VS-6AL chromosomes. These materials could be used for acquiring stable multiple powdery mildew-resistance in wheat breeding programs.
How transposable elements evolve is a key facet in understanding of spontaneous mutation and genomic rearrangements in various organisms. One of the best ways to approach this question is to study a newly evolved transposable element whose presence is restricted to a specific population or strain. The retrotransposons ninja and aurora may provide insights into the process of their evolution, because of their contrasting characteristics, even though they show high sequence identity. The ninja retrotransposon was found in a Drosophila simulans strain in high copy number and is potent in transposition. On the other hand, aurora elements are distributed widely among the species belonging to the Drosophila melanogaster species complex, but are immobile at least in D. melanogaster. In order to distinguish the two closely resembled retrotransposons by molecular means, we determined and compared DNA sequence of the elements, and identified characteristic internal deletions and nucleotide substitutions in 5'-long terminal repeats (LTR). Analyses of the structure of ninja homologs and LTR sequences amplified from both genomic and cloned DNA revealed that the actively transposable ninja elements were present only in D. simulans strains, but inactive aurora elements exist in both D. melanogaster and D. simulans.
The genetic variance of a quantitative trait changes under assortative mating, where the phenotypes of the mates in the trait are correlated. The change is caused by build-up of linkage disequilibria generated due to the genetic similarity between mates. In this paper the change in the genetic variance under assortative mating is investigated using a genetic approach based on the infinitesimal model. Assuming that the genetic effect of each locus is of order n-1/2, where n is the number of loci involved, and formulating the choice of the mate as a selecting function, the linkage disequilibiria are evaluated under repeated cycles of assortative mating. The recursive formula for the genetic variance given by Bulmer (1980), based on the regression of the genetic value of offspring to those of the parents, is obtained by an alternative approach taking the linkage disequilibria into account. This approach can be applied to the case of multiple traits, where two or more traits are simultaneously involved in the assortative mating. A recursive formula for the genetic covariance matrix of multiple traits under assortative mating, which was given by Tallis and Leppard (1987b) based on a statistical approach, is derived using genetic arguments similar to the case of a single trait. It is shown that the equilibrium values of genetic variance of a trait under assortative mating involving the other traits along with the trait are different from those under assortative mating based on the trait alone. In a natural population it is expected that assortative mating is practiced for multiple traits. Thus, multiple traits should be jointly analyzed in research on assortative mating.
We have described a rapid spontaneous conversion in the stationary phase of Escherichia coli strain DOO (crp-) cells as a whole population to crp+ state (Sugino and Morita, 1994). In this paper we have tried to elucidate the molecular basis of this unidirectional conversion by cloning and sequencing of the crp gene in their crp+ and crp- states. We have found that in the original crp- strain, an IS2 element has been inserted between its original promoter and the coding region of the crp gene in the so-called orientation II (Ahmed et al., 1981), accompanied by an 11 bp deletion. Unexpectedly, the crp+ "revertants" derived from the crp- mutant had no difference in sequence from the crp-, either in the coding or the regulatory region. This suggests that a change at another locus, such that this change somehow activates the expression of the crp gene to the level of a normal crp+, is responsible for the apparent reversion from crp- to crp+.