| 1. |
- GUSTAFSSON, L, et al.
(författare)
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LOW GENETIC-VARIATION IN SWEDISH POPULATIONS OF THE RARE SPECIES VICIA-PISIFORMIS (FABACEAE) REVEALED WITH RFLP (RDNA) AND RAPD
- 1994
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Ingår i: Plant Systematics and Evolution. - 0378-2697 .- 1615-6110. ; 189:3-4, s. 133-148
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Tidskriftsartikel (refereegranskat)abstract
- Nine Swedish populations, 1-5 individuals/population, and one cultivated individual of the rare species Vicia pisiformis were investigated for genetic variation. In hybridizations with two rDNA probes using 8 restriction enzymes, only two individuals belonging to one population were polymorphic. A map of the rDNA gene cluster was constructed for four of the restriction enzymes used. Two of the polymorphic sites were mapped and were found to be located outside regions coding for rRNA, presumably caused by single point mutations or small deletions. The repeat length of the rDNA region was c. 10,000 bp, which corresponds well with the size found for other species belonging to Fabaceae. No length polymorphism was found in the intergenic spacer, contrary to the situation found for most other plant species investigated for rDNA variation. The haplotype diversity for the species (Hsp Shannon) was very low (0.055). Within-population values (Hpop) was 0 for all populations except the variable one, which had 0.301. PCR amplification with 6 random primers also revealed very low levels of genetic diversity. A polymorphism was observed in a limited number of individuals for four populations. Hsp was 0.065 and HpopBAR was 0.050. The average D value (Wetton) for the PCR haplotypes was 0.99.
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| 2. |
- Bhalerao, RP, et al.
(författare)
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Cloning of the cpce and cpcf genes from synechococcus sp pcc-6301 and their inactivation in synechococcus sp pcc-7942
- 1994
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Ingår i: Plant Molecular Biology. - 0167-4412 .- 1573-5028. ; 26:1, s. 313-326
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Tidskriftsartikel (refereegranskat)abstract
- Two open reading frames denoted as cpcE and cpcF were cloned and sequenced from Synechococcus sp. PCC 6301. The cpcE and cpcF genes are located downstream of the cpcB2A2 gene cluster in the phycobilisome rod operon and can be transcribed independently of the upstream cpcB2A2 gene cluster. The cpcE and cpcF genes were separately inactivated by insertion of a kanamycin resistance cassette in Synechococcus sp. PCC 7942 to generate mutants R2EKM and R2FKM, respectively, both of which display a substantial reduction in spectroscopically detectable phycocyanin. The levels of beta- and alpha-phycocyanin polypeptides were reduced in the R2EKM and R2FKM mutants although the phycocyanin and linker genes are transcribed at normal levels in the mutants as in the wild type indicating the requirement of the functional cpcE and cpcF genes for normal accumulation of phycocyanin. Two biliprotein fractions were isolated on sucrose density gradient from the R2EKM/R2FKM mutants. The faster sedimenting fraction consisted of intact phycobilisomes. The slower sedimenting biliprotein fraction was found to lack phycocyanin polypeptides, thus no free phycocyanin was detected in the mutants. Characterization of the phycocyanin from the mutants revealed that it was chromophorylated, had a lambda(max) similar to that from the wild type and could be assembled into the phycobilisome rods. Thus, although phycocyanin levels are reduced in the R2EKM and R2FKM mutants, the remaining phycocyanin seems to be chromophorylated and similar to that in the wild type with respect to phycobilisome rod assembly and energy transfer to the core.
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| 3. |
- Bhalerao, RP, et al.
(författare)
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Cloning of the phycobilisome rod linker genes from the cyanobacterium synechococcus sp pcc-6301 och their inactivation in synechococcus sp pcc-7942
- 1993
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Ingår i: Molecular General Genetics. - 0026-8925 .- 1432-1874. ; 237:1-2, s. 89-96
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Tidskriftsartikel (refereegranskat)abstract
- The phycobilisome rod linker genes in the two closely related cyanobacteria Synechococcus sp. PCC 6301 and Synechococcus sp. PCC 7942 were studied. Southern blot analysis showed that the genetic organization of the phycobilisome rod operon is very similar in the two strains. The phycocyanin gene pair is duplicated and separated by a region of about 2.5 kb. The intervening region between the duplicated phycocyanin gene pair was cloned from Synechococcus sp. PCC 6301 and sequenced. Analysis of this DNA sequence revealed the presence of three open reading frames corresponding to 273, 289 and 81 amino acids, respectively. Insertion of a kanamycin resistance cassette into these open reading frames indicated that they corresponded to the genes encoding the 30, 33 and 9 kDa rod linkers, respectively, as judged by the loss of specific linkers from the phycobilisomes of the insertional mutants. Amino acid compositions of the 30 and 33 kDa linkers derived from the DNA sequence were found to deviate from those of purified 33 and 30 kDa linkers in the amounts of glutamic acid/glutamine residues. On the basis of similarity of the amino acid sequence of the rod linkers between Synechococcus sp. PCC 6301 and Calothrix sp. PCC 7601 we name the genes encoding the 30, 33 and 9 kDa linkers cpcH, cpcI and cpcD, respectively. The three linker genes were found to be co-transcribed on an mRNA of 3700 nucleotides. However, we also detected a smaller species of mRNA, of 3400 nucleotides, which would encode only the cpcH and cpcI genes. The 30 kDa linker was still found in phycobilisome rods lacking the 33 kDa linker and the 9 kDa linker was detected in mutants lacking the 33 or the 30 kDa linkers. Free phycocyanin was found in the mutants lacking the 33 or the 30 kDa linkers, whereas no free phycocyanin could be found in the mutant lacking the 9 kDa linker.
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| 4. |
- Bhalerao, RP, et al.
(författare)
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Factors influencing the phycobilisome rod composition of the cyanobacterium synechococcus sp pcc-7942 : effects of reduced phycocyanin content, lack of rod-linkers, and over-expression of the rod-terminating linker
- 1994
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Ingår i: Physiologia Plantarum. - 0031-9317 .- 1399-3054. ; 90:1, s. 187-197
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Tidskriftsartikel (refereegranskat)abstract
- Four novel mutants with altered phycobilisomes were constructed in the cyanobacterium Synechococcus 7942 to study factors influencing the rod length and composition. These mutants show (1) reduced phycocyanin content, (2) reduced phycocyanin content combined with loss of the 33 kDa linker, (3) loss of the 30 kDa rod-linker and (4) overexpression of the 9 kDa rod terminating linker. For these mutants we determined the 33 to 27 kDa and 30 to 27 kDa linker ratios in the isolated phycobilisomes and compared these ratios with those in the wild type. The 30 kDa linker can be incorporated into the rods in absence of the 33 kDa linker. The incorporation of the 30 kDa linker is lower in absence of the 33 kDa linker. When the 30 kDa linker is missing, an increase in the level of the 33 kDa linker is seen, indicating that there could be an excess of the 33 kDa linker in the cells. Our results also show that a reduction in the phycocyanin content causes a decrease in the rod length simultaneously with a reduction of the 30/27 linker ratio, without altering the 33/27 ratio. Reduced phycocyanin content and absence of the 33 kDa linker cause a dramatic reduction in the incorporation of the 30 kDa linker into the rods in the mutant B2SMIKM. Over-expression of the 9 kDa linker results in a decreased incorporation of both the 33 and 30 kDa linkers into the rods, the effect being more pronounced for the 30 kDa linker. This result indicates that the level of the 9 kDa linker relative to those of the 33 and the 30 kDa linkers may be an important determinant of the phycobilisome rod length.
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| 5. |
- Bhalerao, RP, et al.
(författare)
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Structure and energy-transfer of the phycobilisome in a linker protein replacement mutant of cyanobacterium synechococcus-7942
- 1991
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Ingår i: Biochimica et Biophysica Acta. - 0006-3002 .- 1878-2434. ; 1060:1, s. 59-66
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Tidskriftsartikel (refereegranskat)abstract
- The role of the linker proteins in the biogenesis and energy transfer of the phycobilisome rod was monitored by making insertional inactivation in the cpcI gene coding for the core-proximal 33 kilodalton (kDa) protein in the cyanobacterium Synechococcus 7942. The insertion leaves the cpcH gene coding for the core-distal 30 kDa protein intact and functional. Analysis of the phycobilisome protein composition of the cpcI mutant shows that the 30 kDa protein is present in normal amounts in the rod, indicating that the 30 kDa linker protein can replace the 33 kDa protein in the biogenesis and structural integrity of the rod. The absorption and fluorescence characteristics of the mutated phycobilisome is almost indistinguishable from that of the wild-type of the same rod length. The fluorescence kinetics from the cpcI mutant show that the dominating decay component has a lifetime from phycocyanin of 69 ps as compared to 72 ps found for the wild-type phycobilisome with the same rod length. The results show that replacing the 33 kDa for the 30 kDa linker in the rod does not alter the energy harvesting or the energy transfer characteristics of the rod in contrast to what has been concluded from data obtained from in vitro experiments. We conclude that the linker polypeptides have only a minor influence on the energy transfer characteristics of the rod but are mainly involved in determining the length of the rod in response to changing environmental light conditions.
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| 6. |
- CLARKE, AK, et al.
(författare)
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2 FUNCTIONALLY DISTINCT FORMS OF THE PHOTOSYSTEM-II REACTION-CENTER PROTEIN D1 IN THE CYANOBACTERIUM SYNECHOCOCCUS SP PCC 7942
- 1993
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 90:24, s. 11985-11989
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Tidskriftsartikel (refereegranskat)abstract
- The cyanobacterium Synechococcus sp. PCC 7942 possesses a small psbA multigene family that codes for two distinct forms of the photosystem II reaction-center protein D1 (D1:1 and D1:2). We showed previously that the normally predominant D1 form (D1:1) was rapidly replaced with the alternative D1:2 when cells adapted to a photon irradiance of 50 mumol/m-2.s-1 are shifted to 500 mumol.m-2.s-1 and that this interchange was readily reversible once cells were allowed to recover under the original growth conditions. By using the psbA inactivation mutants R2S2C3 and R2K1 (which synthesize only D1:1 and D1:2, respectively), we showed that this interchange between D1 forms was essential for limiting the degree of photoinhibition as well as enabling a rapid recovery of photosynthesis. In this report, we have extended these findings by examining whether any intrinsic functional differences exist between the two D1 forms that may afford increased resistance to photoinhibition. Initial studies on the rate of D1 degradation at three photon-irradiances (50, 200, and 500 mumol.m-2.s-1) showed that the rates of degradation for both D1 forms increase with increasing photon flux density but that there was no significant difference between D1:1 and D1:2. Analysis of light-response curves for oxygen evolution for the mutants R2S2C3 and R2K1 revealed that cells with photosystem II reaction centers containing D1:2 have a higher apparent quantum yield (almost-equal-to 25%) than cells possessing D1:1. Further studies using chlorophyll a fluorescence measurements confirmed that R2K1 has a higher photochemical yield than R2S2C3; that is, a more efficient conversion of excitation energy from photon absorption into photochemistry. We believe that the higher photochemical efficiency of reaction centers containing D1:2 is causally related to the preferential induction of D1:2 at high light and thus may be an integral component of the protection mechanism within Synechococcus sp. PCC 7942 against photoinhibition.
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| 7. |
- CLARKE, AK, et al.
(författare)
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IDENTIFICATION AND EXPRESSION OF THE CHLOROPLAST CLPP GENE IN THE CONIFER PINUS-CONTORTA
- 1994
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Ingår i: Plant Molecular Biology. - 0167-4412 .- 1573-5028. ; 26:3, s. 851-862
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Tidskriftsartikel (refereegranskat)abstract
- The clpP gene from the conifer Pinus contorta was identified and isolated from a chloroplast genomic library by heterologous hybridisation to the second exon of the chloroplast clpP gene in tobacco. DNA sequencing of two overlapping clones revealed an uninterrupted 615 bp open-reading frame with 41 to 65% similarity to the clpP genes in five other chloroplast genomes and Escherichia coli. The 615 bp sequence in P. contorta contained perfectly matched motifs for the serine and histidine active sites of the GlpP protease in E. coli. The location of the clpP gene was determined using a physical map of the P. contorta chloroplast genome, and was found to lie within a 10 kb region between the psbE/F and vpoB genes. Sequencing of the regions adjacent to the clpP gene revealed the first exon of the rps12 gene located 135 bp downstream. The genomic position of the first exon of the rps12 gene in relation to the clpP gene is conserved for all other chloroplast clpP genes identified so far. Northern blot analysis showed that the clpP gene in both P. contorta and P. sylvestris was present in several transcript of different length, ranging from 0.8 to 2.4 kb. The two longer transcripts in P. contorta also included the first exon of the rps12 gene. Mapping of the 5' end of the clpP transcripts by primer extension, however, revealed a single transcription initiation site 53 bp upstream of the first ATG codon. Analysis of total RNA isolated from The two pine species grown in darkness or moderate light conditions (250 mu mol photons m(-2) s(-1)) showed no significant difference in the level of expression of the clpP gene. The results suggest that the clpP gene in conifers is part of an operon which includes the first exon of the rps12 and the entire rp120 gene, and is expressed in a light-independent manner as a polycistronic precursor which later undergoes post-transcriptional processing to give the mature monocistronic clpP mRNA.
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| 8. |
- CLARKE, AK, et al.
(författare)
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RAPID INTERCHANGE BETWEEN 2 DISTINCT FORMS OF CYANOBACTERIAL PHOTOSYSTEM-II REACTION-CENTER PROTEIN-D1 IN RESPONSE TO PHOTOINHIBITION
- 1993
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 90:21, s. 9973-9977
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Tidskriftsartikel (refereegranskat)abstract
- We have studied photoinhibition of photosynthesis in the cyanobacterium Synechococcus sp. PCC 7942, which possesses two distinct forms of the photosystem II reaction-center protein D1 (D1:1 and D1:2). We report here that when cells adapted to a growth irradiance of 50 mumol.m-2.s-1 are exposed to an irradiance of 500 mumol.m-2.s-1, the normally predominant D1 form (D1:1) is rapidly replaced with the alternative D1:2. This interchange is not only complete within the first hour of photoinhibition but is also fully reversible once cells are returned to 50 mumol.m-2.s-1. By using a mutant that synthesizes only D1:1, we show that the failure to replace D1:1 with D1:2 during photoinhibition results in severe loss of photosynthetic activity as well as a diminished capacity to recover after the stress period. We believe that this interchange between D1 forms may constitute an active component in a protection mechanism unique among photosynthetic organisms that enables cyanobacteria to effectively cope with and recover from photoinhibition.
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| 9. |
- Gustafsson, Petter, 1948-, et al.
(författare)
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STRUCTURE AND REGULATION OF PHOTOSYNTHESIS GENES IN PINUS-SYLVESTRIS (SCOTS PINE) AND PINUS-CONTORTA (LODGEPOLE PINE)
- 1991
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Ingår i: Forest Ecology and Management. - 0378-1127 .- 1872-7042. ; 43:3-4, s. 287-300
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Tidskriftsartikel (refereegranskat)abstract
- The structure and regulation of one nuclear and one chloroplast gene was studied in Pinus sylvestris (Scots pine) and Pinus contorta (lodgepole pine). cDNA copies of the nuclear located cab genes of Pinus sylvestris, coding for the light-harvesting chlorophyll a/b-binding proteins of photosystem II (LHC-II), were cloned. cab-II genes coding for both types of LHC-II polypeptides, Types 1 and 2, were found. An analysis of the DNA sequences of several different cab-II cDNAs shows that they have a high bias for the nucleotides G and C at the third base positions of the codons, making them more similar to monocot than to dicot genes. Two of the three genes were found to be located within CpG islands. The cab-II genes were found to be expressed in dark-grown seedlings in contrast to what has been found for most angiosperms. The chloroplast genomes of conifers were shown to lack the inverted repeat organization normally found in higher plants, mosses and green algae. The psbA gene, located in the chloroplast genome and coding for the D1 polypeptide in the reaction center of photosystem II, was found to be tandemly duplicated in P. contorta. Cloning and sequence analysis of the two psbA genes and the surrounding regions showed that the duplicated segment is 1.97 kb long and that it ends 19 bp downstream from the psbA stop codon. The corresponding locus of P. sylvestris, which lacks the duplication, was cloned and characterized. A comparison with P. contorta indicates how the duplication/insertion event has occurred. A comparison of third codon position between P. contorta psbA and that of other plants indicated an almost equidistant evolutionary relationship between P. contorta, spinach (or barley) and Marchantia polymorpha.
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| 10. |
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