Intergenic Recombination Research Articles

The role of bacteriophage T7 exonuclease (gene 6) in genetic recombination and production of concatemers

Genetic analysis reported here shows that bacteriophage T7 exonuclease (gene 6) is necessary for intragenic and intergenic recombination in several areas of the T7 genetic map. This supports our previous conclusion ( Lee & Miller, 1974) that the enzyme is necessary for T7 molecular recombination. Results of sucrose gradient analysis show that DNA concatemers are formed when both the T7 exonuclease (gene 6) and the T7 endonuclease (gene 3) are absent. Further results show that concatemers cannot be maintained in the absence of the exonuclease unless the endonuclease is also eliminated. Therefore, concatemers are formed by a process other than normal phage recombination. Selective defects in the recombination system do interfere with the stability of concatemers, however.

Influence of edeine on intergenic and interallelic recombination in Neurospora crassa

The effect of edeine and the mutation edr-2 to edeine resistance on genetic recombination in Neurospora crassa was investigated. For this purpose crosses between pairs of edeine sensitive and edeine resistant strains respectively were set up without or in the presence of the drug (0-750 microg/ml). The genetic markers ylo-1, ad-1, pan-2 (B3 and B5) and tryp-2, all on linkage group VI, were used for scoring recombinants. These were ad+, tryp+ (intergenic recombination) and pan+ (interallelic recombination). Frequencies of about 6-7% for intergenic and of about 0.4% for interallelic recombination were found in crosses between eds strains and ed(r) strains respectively, if edeine was absent. However, crosses in the presence of edeine gave higher frequencies of both intergenic and interallelic recombination (about 12% intergenic and 1% interallelic with 180 to 200 microg ed/ml). The pan+ prototrophs (interallelic recombinants) obtained in the different crosses were tested for distribution of outside markers. The data thus obtained revealed that under the effect of both the mutation to edeine resistance and edeine itself the relative number of noncrossover (gene conversion) recombinants decreases in favour of crossover recombinants, and the relative number of double crossover recombinants (events outside the pan locus) decreases in favour of single crossover recombinants. It is concluded that a) edeine and the mutation ed(r)-2 to edeine resistance affect recombination via related pathways, and b) noncrossover and crossover recombinants are caused by different molecular mechanisms, in agreement with the work of other authors.

Genetic effects of formaldehyde in yeast. I. Influence of the growth stages on killing and recombination

In random cultures, stationary phase cells of Saccharomyces cerevisieae are more resistant to killing induced by formaldehyde than are exponentially growing cells. It is shown that this compound induces intra- and intergenic recombination in this eucaryotic organism. In synchronized populations the lag and G1 phases demonstrate the higher resistance to boFth killing and induction of recombinants by formaldehyde whereas maximal sensitivity occurs during the end of G2 and/or the mitotic division. This pattern is in contrast with that found after treatments by ionizing or ultraviolet radiations.

Intragenic recombination at the white locus of Drosophila hydei.

A fine-structure analysis of the white locus in Drosophila hydei was carried out by means of allele recombination. Four mutants, derived from wild type, mapped at three subloci. These are possibly homologous to the main subloci 2, 3, and 4 of D. melanogaster. Three secondary mutants, derived from the primary wiv allele, were located in the proximal part of the gene. One of them appeared as a homoallele of the original wiv, whereas the remaining two are better explained either as double mutants or as mutants which facilitate irregular exchange. Intragenic recombination at the white locus seems to be more frequent in D. hydei than in D. melanogaster. The comparatively high incidence is probably a general characteristic, common to intragenic and intergenic recombination in D. hydei.

Mating type and sporulation in yeast. II. Meiosis, recombination, and radiation sensitivity in an alpha-alpha diploid with altered sporulation control.

In wild-type S. cerevisiae, diploid cells must be heterozygous at the mating-type locus in order to sporulate. In the preceding paper, we described a number of mutants (CSP mutants), isolated from nonsporulating aa and alpha-alpha parent strains, in which sporulation appeared to be uncoupled from control by mating type. The characterization of one of these mutants (CSP1) is now extended to other processes controlled by mating type. This mutant is indistinguishable from alpha-alpha cells and unlike aalpha cells for mating factor production and response, zygote formation, intragenic mitotic recombination, and for X-ray sensitivity. The mutant apparently undergoes a full round of DNA synthesis in sporulation medium, but with delayed kinetics. Only 20% of the cells complete sporulation. Among spores in completed asci, the frequency of both intra- and intergenic recombination is the same as it is for spores produced by aalpha cells. However, experiments in which cells were shifted from sporulation medium back to minimal growth medium gave a frequency of meiotic recombination between ade2 or leu2 heteroalleles only 25% to 29% as high for CSP1 alpha-alpha diploid or CSP1 aa disomic cells as for aalpha diploid or disomic cells. Because the latter result, indicating recombination defectiveness, measured recombinant production in the entire cell population, whereas the result indicating normal recombination sampled only completed spores, we infer that all meiotic recombination events occuring in the population of CSP1 alpha-alpha cells are concentrated in those few cells which complete sporulation. This high degree of correlation between meiotic recombination and the completion of meiosis and sporulation suggests that recombination may be required for proper meiotic chromosome segregation in yeast just as it appears to be in maize and in Drosophila.

Isolation and characterization of mutator strains of Escherichia coli K-12.

A selection procedure was devised to select for mutants of Escherichia coli K-12 with enhanced rates of spontaneous frameshift mutation. Three types of mutants were isolated. Two of the mutations apparently represent alleles of previously isolated mutL13 and mutS3. The third type of mutation, represented by two alleles, lies between lysA and thyA, and has been designated mutR. mutR increases the rate of spontaneous frameshift mutation and also the rate of base substitution mutations. The mutator phenotype is recessive. Reversion of a lac amber mutation located on an episome is increased in the presence of the mutator, indicating that mutR can act in trans. No change in sensitivity to ultraviolet irradiation or mitomycin C could be found when mutR34 was compared to the isogenic mutR+ strain. The mutator's activity was little affected by the type of medium in which the strain was grown. Deoxyribonucleoside triphosphate pools were normal in mutR34. Intergenic recombination frequencies were the same in mutR and mutR and mutR+ strains, but a two- to threefold increase in intragenic recombination was observed in Hfr times Fminus crosses when the recipeint was mutR34 as compared with mutR+. This increase appeared independent of the distance between the two markers within the gene in which the crossover took place.

The suppression of gene conversion and intragenic crossing over in Ascobolus immersus: evidence for modifiers acting in the heterozygous state.

Four distinct systems cv1, cv2, cv4 and cv6 behaving as monogenic factors have been found in the stock 28 of Ascobolus immersus. They each specifically suppress gene conversion in a spore pigmentation gene (respectively, b1, b2, b4 and b6). In the gene b2, where this phenomenon was studied in most detail, the effect of cv2 on the suppression of gene conversion is almost total; intragenic recombination is very strongly diminished and crossing over is also affected. In the genes b1, b2 and b4 the suppression of conversion is observed only when the corresponding modifier (cv1, cv2 and cv4) is in the heterozygous condition. It is likely that cv6 acts in the same way on b6. cv1 and b1 on one hand, cv2 and b2 on the other, are very closely linked; no recombination was observed between these modifiers and the corresponding spore pigmentation genes. The same genetic distance is observed between the linked genes b4 and b6 and between cv4 and cv6. This could indicate that these two modifiers are also closely linked to the corresponding genes. A possible effect of cv4 on intergenic recombination between b4 and b6 was shown: here again it acts by diminishing the frequency of recombination when cv4 is in the heterozygous state. The problem of the nature of the cv modifiers is discussed.

Mutations affecting meiosis in Podospora anserina. II. Effect of mei2 mutants on recombination.

Three meiosis-deficient mutants of gene mei2 (mei2-1, mei-2-2 and mei2-3) are blocked during the prophase I of meiosis, before normal pachytene. The mutant mei-2-2 is leaky and there is a partial complementation in crosses mei2-2xmei-2-1 and mei2-2xmei2-3. It has thus been possible to analyse descendants of these crosses. This analysis shows an important alteration in recombination frequencies on at least three different linkage groups. Recombination frequencies appear to be increased near the centromere and decreased in other regions of the chromosomes. This coincides with a decrease in chiasma interference. Intergenic recombination is increased in a locus located very near to the chromosome II centromere. Moreover, the relative proportion of crossovers among the recombination events is stronger than in the control. Though it is impossible at present to formulate a precise hypothesis for the action of the mei2 gene at the molecular level, it is proposed that it might well control a stage of the DNA repair or synthesis.

Recombination during blocked chromosome replication in temperature-sensitive strains ofUstilago maydis

SUMMARYtsd1–1 strains ofUstilago maydisare blocked in DNA synthesis under restrictive conditions. On recovery from the block, both intergenic and allelic recombination are enhanced, and aneuploids are generated. UV is synergistic in the induction of recombination. Direct assays of allelic recombination have shown that gene conversion occurs during the block in DNA synthesis.

The genetic implications of u.v. light exposure and liquid-holding post-treatment in the yeastSaccharomyces cerevisiae

SUMMARYAfter liquid-holding treatment in saline, cultures of yeast exposed to u.v. light showed an increased resistance to a second exposure to u.v. irradiation. This increase in resistance during a second series of u.v. exposures was correlated with an increase in the induction of intragenic recombinants. In contrast, no increase in the frequency of the intergenic recombinants or mutations to prototrophy could be detected during the second u.v. dose range. The results obtained could not be explained by the induction of meiosis during liquid holding or by changes in the timing of cell division after u.v. exposure.A model of u.v. repair in yeast is postulated in which liquid-holding treatment results in changes in the proportions of lesions repaired by excision repair and recombination repair (respectively).

Mutations affecting meiosis in Podospora anserina. I. Cytological studies.

Three meiosis-deficient mutants of gene mei2 (mei2-1, mei-2-2 and mei2-3) are blocked during the prophase I of meiosis, before normal pachytene. The mutant mei-2-2 is leaky and there is a partial complementation in crosses mei2-2xmei-2-1 and mei2-2xmei2-3. It has thus been possible to analyse descendants of these crosses. This analysis shows an important alteration in recombination frequencies on at least three different linkage groups. Recombination frequencies appear to be increased near the centromere and decreased in other regions of the chromosomes. This coincides with a decrease in chiasma interference. Intergenic recombination is increased in a locus located very near to the chromosome II centromere. Moreover, the relative proportion of crossovers among the recombination events is stronger than in the control. Though it is impossible at present to formulate a precise hypothesis for the action of the mei2 gene at the molecular level, it is proposed that it might well control a stage of the DNA repair or synthesis.

Radiation-Induced Recombination in Saccharomyces: The Genetic Control of Recombination in Mitosis and Meiosis

Four rec genes, responsible for a deficient mitotic conversion in yeast, have been studied for their effects on mitotic crossing over and in meiotic intragenic and intergenic exchanges. Mutations in rec1 do not affect either mitotic segregation of ade2 or meiotic intragenic or intergenic recombination. Homozygous rec2/rec2 diploids yield 1-2% sporulation and ten percent spore viability. They also exhibit normal frequencies of x-ray induced mitotic crossing over of ade2. Mutations in rec3 suppress sporulation, but do not affect mitotic segregation of cyh2. Diploids homozygous for rec4 show normal frequencies for x-ray induced mitotic crossing over in ade8. Such diploids exhibit 60% sporulation and 90% spore viability. Intragenic meiotic recombination between alleles 2 and 17 in arg4 is reduced about 500 times. Meiotic crossing over appears normal for the region between mating type and ade8 and their respective centromers and also in the intervals between thr1 and CUP1 and the centromere of chromosome VIII....

On the effect of caffeine on mutation and recombination in Schizosaccharomyces pombe.

In Schizosaccharomyces pombe the experiments performed in order to assess some possible influence of caffeine on biological processes related to the induced or spontaneous mutations and meiotic recombinations, have shown that caffeine decreases UV- and NG-induced forward mutations at ad-6 and ad-7 loci and UV-induced reverse-mutations of his- mutants whereas spontaneous mutations to adenine or histidine independence were not affected; intergenic meiotic recombinations in the MT chromosomal region were also decreased when caffeine was present in the crossing medium.

Ultraviolet-light sensitive mutants of Aspergillus nidulans

14 mutants of Aspergillus nidulans were isolated which were more sensitive to the lethal effects of UV light than wild-type strains. Haploidization analysis of 6 mutants showed that 4 are on linkage group IV, 1 on V and the other on II or VIII. The sensitivity of 6 mutants to nitrous acid was tested and it was found that ranking in terms of UV sensitivity paralleled ranking in terms of nitrous acid sensitivity. No effects on the frequency of meiotic intergenic recombination were found in crosses heterozygous for 7 of the mutants. 1 of these mutants was tested in a homozygous cross and again no effects were observed. The mutations were inherited as single mendelian genes and the survival of diploid conidia after UV irradiation suggested that they were also recessive. 5 mutants had an effect on ascospore development. Viable ascospores were recovered from crosses of these mutant strains in reduced numbers.

Comparison of the effects of ultraviolet light and ethylmethanesulphonate upon the frequency of mitotic recombination in yeast.

Diploid cultures of the yeast Saccharomyces cerevisiae were treated with UV light and ethylmethanesulphonate and scored for mitotic recombination and cell survival. Intergenic recombination was induced to the same frequency by both treatments at the same survival level whereas EMS induced more intragenic recombinants than UV treatment at the same survival.

Recombination in ultraviolet-sensitive strains of Saccharomyces cerevisiae

Intergenic and intragenic recombination during vegetative growth and at meiosis has been studied by selective plating techniques in diploid strains of yeast homozygous for alleles of six genes governing sensitivity to ultraviolet radiation. The following results were obtained: 1. ( 1) None of the uvs mutations caused a significant increase in spontaneous mitotic intragenic recombination. 2. ( 2)The frequency of intragenic mitotic recombination (gene conversion) induced by UV light was greater among survivors in the sensitive strains compared to wild-type controls, but was less when compared in terms of equal survival. 3. ( 3) The dose-response curves for UV-induced mitotic recombination between the locus of a gene and its centromere were complex, but in most cases the uvs diploids showed a greater response than the controls. 4. ( 4) No differences from the controls were found in the frequency of intragenic or intergenic recombination at meiosis, but significant differences were found between certain uvs strains.

The effects of dark holding and photoreactivation on ultraviolet light-induced mitotic recombination and survival in yeast.

UV induces lesions in DNA which lead to the death of cells, mutation and, in yeast, intragenic and intergenic mitotic recombination. We have investigated the interaction of two post-treatments, dark holding and photoreactivation, on the frequencies of these events. It was found that dark holding reduces cell death and intergenic recombinants, but causes an increase in intragenic recombination frequency. Photoreactivation reduces cell death and intragenic recombination, but has no effect on intergenic recombination. After dark holding, photoreactivation has no further effect on cell survival or intergenic recombination, but may reduce the frequency of intragenic recombinants. After photoreactivation, dark holding still causes an increase in cell survival and the frequency of intragenic recombination, and reduces the frequency of intergenic recombination.

Mitotic recombination induced by ultraviolet light in synchronous cultures of yeast

Samples taken at intervals from a synchronised mitotic diploid culture of Saccharomyces cerevisiae were treated with ultraviolet light and with photoreactivating white light. They were scored for survival and for the incidence of intergenic and intragenic recombination. It was found that the frequency of all three events varied through the cycle, cell death and intergenic recombination reaching peaks during DNA synthesis and intragenic recombination at the end of DNA synthesis. The periods of maximum photoreactivation coincided with periods of maximum induction of a given event by the ultraviolet light. While cell death showed some photoreactivation at all stages of the cycle, both kinds of UV-induced recombination at some stages responded hardly or not all to photoreactivating light. The interpretation of the results is discussed.

The mechanism of recombination in Chlamydomonas reinhardi II. The influence of inhibitors of DNA synthesis on intergenic recombination

The mechanism of recombination in Chlamydomonas reinhardi II. The influence of inhibitors of DNA synthesis on intergenic recombination

The mechanism of recombination in Chlamydomonas reinhardi I. The influence of inhibitors of protein synthesis on intergenic recombination

Abstract Germinating zygospores of Chlamydomonas reinhardi were exposed to inhibitors of DNA synthesis, at various stages of the pre-meiotic interphase and of meiosis, and effects on recombination in the arg 1-pab 2 region of linkage group I studied. Different inhibitors evoked different patterns of response although the responsive periods were similar; these were confined to two developmental stages—one premeiosis, probably at the time of DNA synthesis and the other in pachytene. The results indicate that the primary events of inter-genic recombination occur in DNA molecules and that some degree of DNA synthesis is associated with the process. Furthermore it can be concluded that the terminal events of crossing-over occur after the main period of DNA synthesis, probably at some stage in pachytene.