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131.
A critical stage in pollen development is the dissolution of the four products of meiosis, the tetrads, into free microspores. The tetrads are surrounded by a thick callose wall composed of -1,3-glucan. At the completion of meiosis, the tetrads are released into the anther locule after hydrolysis of the callose by a -1,3-glucanase. Using the polymerase chain reaction, we have amplified and subsequently cloned a cDNA corresponding to a -1,3-glucanase, tobacco (Nicotiana tabacum cv. Samsun) anther glucanase (Tag 1), which is expressed exclusively in anthers from meiosis to the free microspore stage of pollen development. The identity of the clone was determined by DNA and deduced protein sequence similarity to other known -1,3-glucanases. Several regions strictly conserved among four classes of glucanases are also conserved in the Tag 1 protein. Tag 1 represents a novel class of -1,3-glucanase based on phylogenetic analysis and RNA expression pattern. Tag 1 RNA was detected in situ only in the tapetum, with maximal expression just prior to tetrad dissolution. Due to its expression pattern and sequence similarity to other -1,3-glucanases, we believe Tag 1 may be involved in tetrad dissolution. 相似文献
132.
Hanns Ulrich Seitz Dorothea Elisabeth Gärtner 《Plant Cell, Tissue and Organ Culture》1994,38(2-3):337-344
In contrast to undifferentiated cell suspension cultures of Digitalis lanata, photomixotrophic shoot cultures of Digitalis purpurea accumulate cardiac glycosides in substantial concentrations. They are used to investigate enzymes of the cardenolide pathway. All cardenolides are 5-configurated. The progesterone 5-reductase and the 3-hydroxysteroid-5-oxidoreductase are present in shoot cultures but not in undifferentiated cell cultures. These enzymes provide precursors for cardenolides, whereas the presence of the progesterone 5-reductase, also present in shoot cultures, is discussed with regard to its role in phytosterol biosynthesis and may be attributed to the general steroid pathway. The progesterone 5-reductase had an activity maximum during the early growth period seven days after onset of cultivation, whereas the corresponding progesterone 5-reductase activity was highest on day 11. The maximum cardenolide accumulation was after 24 days. The enzyme activities present in crude extracts from shoot cultures were characterized with regard to their requirements for NADPH and NADH, pH-optimum, temperature optimum, affinity to their substrates and their localization in the cell. The progesterone 5-reductase was purified 769-fold.Abbreviations DW
dry weight
- FW
fresh weight
- PVP
polyvinylpyrrolidone 相似文献
133.
134.
Drosophila melanogaster acetylcholinesterase: Identification and expression of two mutations responsible for cold- and heat-sensitive phenotypes 总被引:2,自引:0,他引:2
Annick Mutero Jean-Marc Bride Madeleine Pralavorio Didier Fournier 《Molecular & general genetics : MGG》1994,243(6):699-705
Ace
IJ29 and Ac
IJ40 are cold- and heat-sensitive variants of the gene coding for acetylcholinesterase in Drosophila melanogaster. In the homozygous condition, these mutations are lethal when animals are raised at restrictive temperatures, i.e., below 23° C for Ace
IJ29 or above 25° C for Ace
IJ40. The coding regions of the gene in these mutants were sequenced and mutations changing Ser374 to Phe in Ace
IJ29 and Pro75 to Leu in Ace
IJ40 were found. Acetylcholinesterases bearing these mutations were expressed in Xenopus oocytes and we found that these mutations decrease the secretion rate of the protein most probably by affecting its folding. This phenomenon is exacerbated at restrictive temperatures decreasing the amount of secreted acetylcholinesterase below the lethality threshold. In parallel, the substitution of the conserved Asp248 by an Asn residue completely inhibits the activity of the enzyme and its secretion, preventing the correct folding of the protein in a non-conditional manner. 相似文献
135.
136.
Endogenous gibberellins (GAs) were extracted and purified from apical buds of Eucalyptus nitens (Deane and Maid.) Maid. and the cambial region of E. globulus (Labill.). then analysed by capillary gas chromatography-mass spectrometry. GA1 GA19 GA20 and GA29 were identified by full scan mass spectra. Kovats retention indices and high resolution selected ion monitoring. Using deuterated internal standards. GA1 . GA19 . GA20 and putative GA29 and GA53 were quantified in the apical buds, while GA4 . GA8 . GA9 and GA44 were shown to be either absent or present at very low levels. From the cambial region. GA1 and GA20 were quantified at levels of 0.30 ng (g fresh weight)-1 and 8.8 ng (g fresh weight)-1 respectively. These data suggest that the early 13-hydroxylation pathway is the dominant pathway for GA biosynthesis in Eucalyptus . 相似文献
137.
Itamar Barash Alexander Faerman Tamar Ratovitsky Raisa Puzis Margaret Nathan David R. Hurwitz Moshe Shani 《Transgenic research》1994,3(3):141-151
We produced transgenic mice carrying the native sheep -lactoglobulin (BLG) or fusion genes composed of the BLG promoter and human serum albumin (HSA) minigenes. BLG was expressed exclusively in the mammary glands of the virgin and lactating transgenic mice evaluated. In contrast, transgenic females carrying the BLG/HSA fusion constructs also expressed the HSA RNA ectopically in skeletal muscle, kidney, brain, spleen, salivary gland and skin. Ectopic expression of HSA RNA was detected only in strains that express the transgene in the mammary gland. There was no obvious correlation between the level of the HSA RNA expressed in the mammary gland and that found ectopically. In three transgenic strains analysed, the expression of HSA RNA in kidney and skeletal muscle increased during pregnancy and lactation, whereas in the brain HSA expression decreased during lactation in one of the strains. HSA protein was synthesized in skeletal muscle and skin of strain #23 and its level was higher in lactating mice compared with virgin mice. Expression of HSA was also analysed in males and was found to be more stringently controlled than in females of the same strains.In situ hybridization analyses localized the expressed transgene in the skin, kidney, brain and salivary glands of various transgenic strains. Distinct strain-specific and cell-type specific HSA expression patterns were observed in the skin. This is in contrast to the exclusive expression of the HSA transgene in epithelial cells surrounding the alveoli of the mammary gland. Taken together, these results suggest that the absence of sufficient mammary-specific regulatory elements in the BLG promoter sequences and/or the juxtaposition of the BLG promoter with the HSA coding sequences leads to novel tissue- and cell-specific expression in ectopic tissues of transgenic mice. 相似文献
138.
Mohammad B. Rashid Katsuhiko Shirahige Naotake Ogasawara Hiroshi Yoshikawa 《Gene》1994,150(2):213-220
We have analyzed the relationship between autonomously replicating sequence (ARS) structure and function for three ARS (ARS605, ARS607 and ARS609) from chromosome VI of Saccharomyces cerevisiae by systematic XhoI-linker mutation in the ARS consensus sequence (ACS) and flanking sequences. All mutations that encroached upon the ACS destroyed ARS activity. DNA sequences stimulative for ARS function were identified on either side of the ACS of ARS605 and only on the 3'-side of the ACS of ARS607. In ARS609, however, no such stimulative sequences were observed. Base substitutions complementary to the wild-type sequence of those stimulative regions, in ARS605 and ARS607, that did not change the AG of unwinding nor affected ARS activity suggests that these regions have, at least, a function as DNA-unwinding elements (DUE). ARS605, ARS607 and ARS609 DNA are of low AG value and showed hypersensitivity to single-strand-specific nuclease when inserted in negatively supercoiled plasmid. Linker mutations inhibitory for ARS activity (5L11 and 7L14) also caused significant changes in local nucleotide (nt) sensitivity within the ACS and its adjoining regions. Complementary base substitutions, however, did not affect these changes in local nt sensitivity. These results imply that the stimulative regions flanking the ACS are necessary to produce an optimum conformation around the ACS which may be important for full ARS activity. 相似文献
139.
Summary A 3D triple resonance experiment has been designed to provide intraresidual and sequential correlations between amide nitrogens and -carbons in uniformly 13C15N-labeled proteins. In-phase 13C magnetization is transferred to the aliphatic side-chain protons via the side-chain carbons using a CC-TOCSY mixing sequence. Thus, the experiment alleviates the resonance assignment process by providing information about the amino acid type as well as establishing sequential connectivities. Leaving the carbonyl spins untouched throughout the transfer from 13C to 1H leads to E.COSY-type cross peaks, from which the 3JH
co coupling constants can be evaluated. The pulse sequence is applied to oxidized Desulfovibrio vulgaris flavodoxin. 相似文献
140.
非同位素PCR-单链构象多态性技术的建立和应用 总被引:2,自引:0,他引:2
PCR-单链构象多态性技术问世以来,成为研究基因突变的工具,特别是在分子肿瘤学研究中,广泛应用于癌基因,抑癌基因突变的研究,常规PCR-SSCP采用同位素标记PCR产物,测序板电泳分离突变,在操作和费用上有种种局限,文章建立了一种非同位素PCR-SSCP技术;通过不对称PCR获得单链,普通PAGE分离,经银染检出突变,用这种方法,还研究了四株鼻咽癌细胞株CNE1,CNE2,HK1和SUNE1中肿瘤 相似文献