植物生理与分子生物学学报最新文献

Chalcone synthase (CHS) catalyses the first and key regulatory step of flavonoid biosynthetic pathway. A chalcone synthase gene was isolated from Ginkgo biloba leaves using the method of rapid amplification of the cDNA ends (RACE). The full-length cDNA, designated as GbCHS2, is 1,608 bp in length (GenBank accession No. DQ054841) and contains an open reading frame of 1,173 bp encoding a protein of 391 amino acids. Alignment of the predicted amino acid sequence of GbCHS2 has been shown to have high sequence similarity with GbCHS1. All the active sites and active site motifs in GbCHS1 protein were also found in GbCHS2. Correlation analysis between CHS activity and flavonoid accumulation during ginkgo leaf growth indicated that CHS might be the rate-limiting enzyme in the biosynthesis pathway of flavonoids in ginkgo leaves. Results of semi-quantitative RT-PCR analysis showed that flavonoid accumulation paralleled the transcription level of change in chs gene, suggesting chs gene as the specific key gene regulating flavonoid accumulation in ginkgo leaves.

Chloroplasts are a vital group of organelles of plants, yet the molecular mechanisms associated with their division remain poorly understood. Recent studies have revealed that the FtsZ protein, known as a key component in prokaryotic cell division, is involved in chloroplast division process. The NtFtsZ2-1 gene was isolated from Nicotiana tabacum by RT-PCR, and the sense and antisense expression plasmids were used to examine the function of NtFtsZ2-1 gene in transgenic tobacco. Light and confocal observations revealed that the normal chloroplast division process was severely disrupted in transgenic plants with enhanced or reduced expression of NtFtsZ2-1 gene. These chloroplasts were abnormally larger in size and fewer in number compared with that of the wild-type tobacco. But the total chloroplast plan area per mesophyll cell was conserved in sense, antisense and wild type tobaccos. Analyses of electron micrographs and chlorophyll content of different transgenic plants showed that constitutively enhancing or inhibiting the expression of NtFtsZ2-1 gene had no direct influence on the ultrastructure and photosynthetic ability of chloroplasts. Basing on these results, we suggest that NtFtsZ2-1 gene is involved in chloroplast division and expansion; the fluctuation of NtFtsZ2-1 expression level would alter normal chloroplast number and size in plant cells. In addition, the similarities of ultrastructure and photosynthetic ability of chloroplasts among sense, antisense and wild type tobaccos implies that a special mechanism regulate the relationship between chloroplast number and size to maximize photosynthetic rate.

Jerusalem artichoke (Helianthus tuberosus L.) seedlings cultured in sandy media were treated with Hoagland nutrition solution with different concentrations of Cd(NO(3))(2) from 0 to 400 micromol/L. After 50 days' treatment, Cd accumulation, activities of peroxidase (POD, EC 1.11.1.7), superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6) were measured and electrophoretograms of POD isoenzymes were analyzed. The accumulation of Cd in seedlings increased from Cd 50-100 micromol/L, after which further increases in Cd concentration resulted in only small increases in accumulation of Cd in seedlings. MDA content was markedly higher than control values indicating the enhanced membrane lipid peroxidation in roots and leaves. POD activities in leaf and root extracts increased with an increase of Cd concentration from 0 to 50 and 100 micromol/L and then decreased with further increases to 200 and 400 micromol/L. Under moderate Cd level of 50-200 micromol/L, SOD activities in leaf and root extracts increased whereas with a higher Cd level of 400 micromol/L marked inhibitions in enzyme activities were observed. With increase in Cd concentration marked elevations in CAT activities in leaves and roots were observed. Results of electrophoresis show that the alteration of POD isoenzyme was noticeable to Cd and an additional POD isoenzyme LP10 appeared. It is suggested that POD isoenzyme of Jerusalem artichoke seedlings could be used as bioindicator for soil contamination by Cd.

In order to explore the relationship between grain yield and photosynthesis, the yield composition and leaf photosynthetic rate in some super hybrid rices and ordinary hybrid rice 'Shanyou 63' as control were measured in 2000-2005. The results were as follows. (1) The yield levels of the four super hybrid rices, 'Pei'ai 64S/E32', 'P88S/0293', 'Jin23A/611' and 'GD-1S/RB207', were significantly higher, being 108%-120% of 'Shanyou 63'. (2) These super hybrid rices had a better plant type with more erect upper layer leaves and bigger panicles or more spikelets per panicle, being 125%-177% of spikelets Shanyou 63 spikelets. (3) Net photosynthetic rates of these super hybrid rices were significantly higher in the second leaf but not necessarily in the first leaf or flag leaf than those of spikelets Shanyou 63 spikelets. (4) The removal of half flag leaf led to a decline in the seed-setting rate, while the removal of half panicle induced its increase in spikelets GD-1S/RB207 spikelets. Hence, higher yield in these super hybrid rices can be attributed to their bigger panicles, better plant type and higher light use efficiency of their canopies. Raising the photosynthetic capacity of each leaf, especially flag leaf, is the key to overcome the photosynthate-source restriction on grain yield and to make a new breakthrough of yield potential in future breeding of super hybrid rice.

The glucose, fructose and sucrose contents and the activities of invertase, sucrose synthase (SS), sucrose phosphate synthase (SPS), hexokinase (HXK) and fructokinase (FRK) of 'Tochiotome' strawberry aggregate fruit were analyzed at various stages of fruit development. The sugar accumulation exhibited the difference between various parts of the fruit. The sugar content is the highest in the top part of ripe 'Tochiotome' strawberry aggregate fruit, followed by the central part, and the basal part was the lowest. Changes in activity of invertase corresponded to the changes of sugar concentration gradient within various parts of the ripe strawberry aggregate fruit. The SS and SPS activities within various parts of aggregate fruit did not showed obvious change that corresponded to the sugar content. At the stage of fruit maturation, the HXK and FRK activities within various parts of strawberry aggregate fruit, however, exhibited an opposite gradient to sugar content. The basal part of aggregate fruit had the highest activities of HXK and FRK, while the top part had the lowest activities. These results suggested that the higher activity of invertase in the top part of aggregate fruit may play a role for rapid cleavage of sucrose and help to form a sucrose gradient in apoplastic space from the basal part to the top part of strawberry aggregate fruit, thus enhance photosynthate translocation from the basal part to the top part of aggregate fruit, which led to higher sugar accumulation in the top part of fruit. In the basal part of aggregate fruit, the higher activities of hexose-metabolizing enzymes may promote the consumption of hexose, and thus resulted in the lowering of sugar content in the basal part than in the top part of strawberry aggregate fruit.

A number of signal pathways have been found through which abundant calcium-stimulated protein kinase activity in plant is associated with calcium-dependent protein kinases (CDPKs) which act as the calcium sensors mediating numerous responses, including hormone signaling. Basing on previous studies, we made additional functional analysis of the gene AtCPK30 encoding a protein kinase in Arabidopsis. Results of semi-quantitative reverse transcription PCR (RT-PCR) analysis indicated that AtCPK30 was highly expressed in root and induced by ABA, IAA, 2,4-D, GA(3) and 6-BA treatment. The physiological roles of AtCPK30 were studied using a gain-of-function approach. Seedlings of AtCPK30 transgenic lines had longer primary roots than those plants of wild-type at the early stages. Interestingly, when these plants grew on MS lack of Ca(2+) including wild-type and transgenic lines, the roots of transgenic line were more sensitive to calcium, lack of Ca(2+) had less effect on roots of transgenic lines than those of wild-type. Treated with several plant hormones, such as ABA, IAA, GA(3) and 6-BA, the roots of seedlings of transgenic line developed abnormally because they were more sensitive to hormones. Furthermore, NPA relatively less inhibited emergency of lateral roots of transgenic line than those of the wild-type. Green fluorescent protein-CPK30 (GFP-CPK30) fusion protein studies revealed the localization of AtCPK30 to both cell wall and plasma membrane. These results suggest that AtCPK30 acts as the calcium sensor and involved in the hormone-signaling pathways.

A full-length 1,043-base-pair cDNA clone encoding a chloroplast copper/zinc superoxide dismutase (Cu/Zn-SOD) of upland cotton was first isolated by rapid amplification of cDNA ends (RACE) from the leaves of the cotton (Gossypium hirsutum L.) variety 'CRI36'. Nucleotide sequence analysis of the clone revealed that it contained the complete coding sequence of the mature SOD isozyme subunit, along with a 60-amino acid transit peptide at N-terminal. The amino acid sequence predicted from the full-length clone showed 66%-74% homology with the amino acid sequences of Cu/Zn-SOD from several other plants. This gene was found to be expressed in the leaves and stems, but not in roots, flowers, and hypocotyls, indicating that the gene was expressed only in green tissues. Also, its expression was found to be most active at seedling stage and declined gradually in later development stages. Expression of this cotton Cu/Zn-SOD gene by using the pET-21a (+) expression vector in E. coli BL21 (DE3) led to the production of a novel 29 kD polypeptide with SOD enzyme activity, confirming that the cloned cotton Cu/Zn-SOD cDNA was indeed encoding a functioning SOD enzyme.

To use different types of promoters in transgenic rice research, the 1.1 kb 5'-upstream regulation region of one of the tomato (Solanum tuberosum L.) Rubisco small subunit gene, rbcS3A, was cloned and its sequences were confirmed by comparison with the known sequences in GenBank. The cloned rbcS3A promoter was fused to the 5'-upstream of GUS (beta-glucuronidase) coding region in a binary vector, and introduced into an elite japonica rice variety by Agrogacterium-mediated transformation. The integration of the GUS fusion gene into the genome of transgenic rice was confirmed by both PCR and Southern blot analysis. The results of both histochemical staining and quantitative analysis of GUS activity showed that the expression level of GUS fusion gene was significantly stronger in stem, leaf blade and sheath than in other organs of transgenic rice plants, and showed highest in the stem, which implies that the tomato rbcS3A promoter can make tissue-specific, in particular in the stem, expression of foreign genes in transgenic rice. The results present here also demonstrate that light induction had no effect on the expression of the foreign gene when regulated by the tomato rbcS3A promoter in transgenic rice. Our results show that the cloned tomato rbcS3A promoter might be very useful for the expression of target genes in transgenic rice, with particularly high efficiency in stem tissues.

The total proteins of Leymus chinensis were extracted from eight-week-old seedlings grown in the greenhouse by TCA-acetone precipitation method. After the lysis buffer was replaced by the start buffer, proteins were fractionated along the first dimension using chromatofocusing (CF). Subsequently the fractions with pI values between 8.5 and 4.0 collected after the first dimension separation were further fractionated by nonporous silica reverse-phase high-performance liquid chromatography (NPS-RP-HPLC, HPRP). With ProteoVue software the pI/UV map was generated to show the protein expression profile of Leymus chinensis. Some experiments were tested to optimize the fraction procedure. Three different elution gradients were employed to get the optimal chromatogram. Comparison of the protein expression pattern between 2D-PAGE and 2D-LC indicated that 2D-LC was a powerful tool in protein fraction. Reproducibility and veracity of the protein patterns were confirmed in different injections of the same sample. A method to fractionate the total protein of Leymus chinensis shoot with two-dimensional liquid chromatography (2D-LC) was founded.

The level of cytosine methylation induced by cadmium in radish (Raphanus sativus L.) genome was analysed using the technique of methylation-sensitive amplified polymorphism (MSAP). The MSAP ratios in radish seedling exposed to cadmium chloride at the concentration of 50, 250 and 500 mg/L were 37%, 43% and 51%, respectively, and the control was 34%; the full methylation levels (C(m)CGG in double strands) were at 23%, 25% and 27%, respectively, while the control was 22%. The level of increase in MSAP and full methylation indicated that de novo methylation occurred in some 5'-CCGG sites under Cd stress. There was significant positive correlation between increase of total DNA methylation level and CdCl(2) concentration. Four types of MSAP patterns: de novo methylation, de-methylation, atypical pattern and no changes of methylation pattern were identified among CdCl(2) treatments and the control. DNA methylation alteration in plants treated with CdCl(2) was mainly through de novo methylation.