Marker Assisted Transfer of Two Powdery Mildew Resistance Genes PmTb7A.1 and PmTb7A.2 from Triticum boeoticum (Boiss.) to Triticum aestivum (L.)

Powdery mildew (PM), caused by Blumeria graminis f. sp. tritici, is one of the important wheat diseases, worldwide. Two PM resistance genes, designated as PmTb7A.1 and PmTb7A.2, were identified in T. boeoticum acc. pau5088 and mapped on chromosome 7AL approximately 48cM apart. Two resistance gene analogue (RGA)-STS markers Ta7AL-4556232 and 7AL-4426363 were identified to be linked to the PmTb7A.1 and PmTb7A.2, at a distance of 0.6cM and 6.0cM, respectively. In the present study, following marker assisted selection (MAS), the two genes were transferred to T. aestivum using T. durum as bridging species. As many as 12,317 florets of F₁ of the cross T. durum /T. boeoticum were pollinated with T. aestivum lines PBW343-IL and PBW621 to produce 61 and 65 seeds, respectively, of three-way F₁. The resulting F₁s of the cross T. durum/T. boeoticum//T. aestivum were screened with marker flanking both the PM resistance genes PmTb7A.1 and PmTb7A.2 (foreground selection) and the selected plants were backcrossed to generate BC₁F₁. Marker assisted selection was carried both in BC₁F₁ and the BC₂F₁ generations. Introgression of alien chromatin in BC₂F₁ plants varied from 15.4 - 62.9 percent. Out of more than 110 BC₂F₁ plants showing introgression for markers linked to the two PM resistance genes, 40 agronomically desirable plants were selected for background selection for the carrier chromosome to identify the plants with minimum of the alien introgression. Cytological analysis showed that most plants have chromosome number ranging from 40-42. The BC₂F₂ plants homozygous for the two genes have been identified. These will be crossed to generate lines combining both the PM resistance genes but with minimal of the alien introgression. The PM resistance gene PmTb7A.1 maps in a region very close to Sr22, a stem rust resistance gene effective against the race Ug99. Analysis of selected plants with markers linked to Sr22 showed introgression of Sr22 from T. boeoticum in several BC₂F₁ plants. Thus, in addition to PM resistance, these progeny might also carry resistance to stem rust race Ug99.     

A sterospecific synthesis of 7alpha-hydroxycholesterol.

The five step synthesis of 7alpha-hydroxycholesterol utilizes the solvolysis of 7alpha-bromocholesterol benzoate with potassium acetate in acetic acid as the key step in controlling the stereospecificity of the reaction sequence. This reaction yields 7alpha-acetoxycholesterol benzoate with retention of configuration at position seven. The diester is readily reduced with lithium aluminum to 7alpha-hydroxycholesterol.     

Primary gastric lymphoma. A survey of 7 cases

Clinical characteristics and response to therapy of 7 patients with primary non-Hodgkin's lymphoma of the stomach and 1 patient with double malignancy are analysed. All patients but one were gastrectomized and subjected to the chemotherapy (COP, CHOP or leukeran + prednison) programme. In 3 patients (2 with lymphocytic lymphoma and 1 with lymphoplasmocytic lymphoma) no recurrence occurred in the course of the observation time ranging from 18 to 20 months. In one patient with centrocytic lymphoma extra-abdominal recurrence was successfully treated with leukeran + prednison. 4 patients (2 with immunoblastic lymphoma and 1 with lymphoplasmocytic lymphoma) with abdominal or extra-abdominal recurrence died after a period ranging from 4 to 29 months despite more aggressive therapy with CHOP programme. Opinions concerning pathogenesis and treatment strategy of those lymphomas are presented too.     

A Sec7-related protein in Paramecium.

We have cloned and sequenced a SEC7-related gene in Paramecium tetraurelia that contains an open reading frame for 1135 amino acids encoding a 133 kDa protein, PSec7. Sec7, first identified in vesicular trafficking mutants in yeast, and its phylogenetic homologues function as guanine-nucleotide exchange factors for small G-proteins such as ARF (ADP-ribosylation factor). The deduced amino acid sequence in PSec7 for the motifs that form the ARF binding site are more than 70% identical to yeast Sec7 and similarly identical to ARNO, the human ARF exchange factor, with correct positioning of the critical glutamic acid residue within the motif region. Overall, the identity of PSec7 to yeast Sec7 is 32%. The deduced amino acid sequence also has five sequences that resemble IQ motifs, EF hand binding domains found in all myosins, and two pleckstrin homology domains. Similar sequences are present in yeast Sec7 and other Sec7-related molecules. A protein kinase A phosphorylation site may also be present. Southern blots suggest that a single gene encodes PSec7. Northern blots show that the message encoding PSec7 is induced on deciliation, followed by ciliogenesis, which suggests a role for PSec7 in cilia such as transport or targeting of ciliary membrane components.     

锌_7与镉_7－金属硫蛋白清除羟自由基的比较

分离及纯化兔肝金属硫蛋白制备去金属金属硫蛋白、锌７与镉７金属硫蛋白．在不同ｐＨ条件下,比较后二者清除羟自由基能力;在ｐＨ６条件下,比较锌７－金属硫蛋白与有关蛋白和无机锌盐清除羟自由基效果．结论是在近生理ｐＨ条件下锌７－金属硫蛋白清除羟自由基能力远强于镉７－金属硫蛋白．金属硫蛋白清除羟自由基的能力主要来源于蛋白中处于还原态的流基．     

A Chromosomal Study on 7 Species of Smilax L.

The chromosome numbers and karyotypes of 7 species of Smilax L. in Liliaceae (s. 1.) are cytotaxonomically studied in this work. Their karyotypic characters, distinction between the species and the chromosomal basis of sexual differentiation are discussed. The karyotypes of most species are first reported. The results are shown as follows (see Tables 1-4 for the chromosome parameters and the karyotype constitution; Fig. 1 for their idiograms): 1. Smilax nipponica Miq. The species is one of the herbaceous species distributed in East Asia. Two karyotypes, 2n = 26(type A) and 2n = 32 (type B), are found in the species (Plate 1: 1-7). The karyotype of No. 88032 (uncertain of -L--M--S- sexuality) is 2n = 26 = 2m + 6st + 6m + 4sm + 6sm + 2st. The karyotype has 4 pairs of L chromosomes, of which the first three pairs are subterminal, and the 4th is median. The karyotype belongs to 3B. No. 88045 (the male) and No. 88046 (the female) have 2n = 32. Their karyotypes are basically uniform, and both are -L--M-- S 2n=32= 2m+4sm+ 2st+ 2m+4sm+ 6m+ 10sm + 2st, also with 4 pairs of L chromosomes, but the 2nd pair is median, and thus different from the type A. The karyotype belongs to 3B. The first pair of chromosomes of the male are distinctly unequal in length, with the D. V. (0.93) of relative length between them obviously greater than that of the female (0.1). The pair seems to be of sex-chromosomes. Sixteen bivalents (n= 16) were observed at PMCs MI of No. 88045 (Plate 1: 4). The major difference between the karyotypes A and B are greater relative length of L chromosomes in the type A than in the type B, and the increase of chromosome number in the karyotype B mainly due to the increase of st chromosomes. Nakajima (1937)reports 2n= 30 for S. hederacea var. nipponica (=S. nipponica, Wang and Tang, 1980). 2. S. riparia A. DC. This species is also herbaceous, distributed in East Asia. Thirty chromosomes were found in root-tip cells (uncertain of sexuality). The kar -L--M--S-yotype is 2n = 30 = 8st + 6sm + 2st + 6m + 6sm + 2st (Plate 3: 1, 5), consisting mainly of sm and st chromosomes. There are 4 pairs of L chromosomes which are all subterminal and the m chromosomes appear to fall all into S category. Though the karyotype belongs to 3B, it is less symmetrical than that of S. nipponica. The species is karyologically rather different from S. nipponica, therefore. The first pair of chromosomes of this material are unequal in length, and it may be a male. The karyotype of this species is first reported. 3. S. sieboldii Miq. The species is a thorny climbing shrub, distributed in East Asia. At PMCs All, 16 chromosomes (n= 16) were found (Plate 2: 6), in accordance with Nakajima's (1933) report for a Japanese material. 4. S. china L. This species, a thorny climbing shrub, is of a wide distribution range mainly in East Asia and Southeast Asia. Two karyotypes were observed in different populations. (1) The population from Xikou has 2n = 96(6x) = 20st+L- -M- 6t + 6sm + 12st + 52(S) (Plate 3:7), of which the first three pairs of chromosomes are terminal, different from those in the other species. The arm ratios of both L and M chromosomes are larger than 2.0, which resembles those of S. davidiana. (2) PMCs MI of the population from Shangyu shew 15 chromosomes (n 15). The hexaploid of the species is recorded for the first time. Hsu (1967,1971) reported 2n = 30 from Taiwai and Nakajima (1937) recorded n = 30 from Japan, which indicates that the karyotype of the species varies not only in ploidy, but also in number. 5. S. davidiana A. DC. The somatic cells were found to have 32 chromosomes, and PMCs MI shew 16 bivalents (Plate 2: 1-5). The karyotype is 2n = 32=-L- -M- -S 8st + 4sm + 4st + 8sm + 8st. The karyotype belongs to 3B, and is less symmetrical than those in herbaceous species. The D. V. (0.20) of relative length between the two homologues of the first pair is slightly larger in the male than in the female (0.14), and it is thus difficult to determine whether they are sexual chromosomes or not. 6. S. glabra Roxb. The species is a non-thorny climbing shrub, distributed in East Asia and Southeast Asia. 32 chromosomes were found in somatic cells. The -L- -M- - Skaryotype is 2n= 32= 8st + 10st+6sm+8st (Plate 3: 2, 6),with only 3 pairs of sm chromosomes (12, 13 and 16th). The karyotype is more asymmetric than that of S. davidiana, although it is also of 3B (Table 1). The karyotype is first reported for the species. 7. S. nervo-marginata Hay. var. liukiuensis (Hay.) Wang et Tang The variety has a relatively narrow distribution range, mainly occurring in eastern China. The chromosomal number of somatic cells is 2n= 32 (Plate 3: 3-4). The karyotype is -L- -M- -S 2n = 32 = 2sm + 6st + 2sm + 2st + 2m + 6sm + 12st, evidently different from that of S. glabra. The first pair of chromosomes are submedian, and much longer than the 2nd to 4th pairs. The ratio in length of the largest chromosome to the smallest one is 4.3. The symmetric degree is of 3C, a unique type. The karyotype of the species is reported for the first time. In Smilax, the known basic numbers are 13, 15, 16 and 17. The two herbaceous species distributed in East Asia have three basic numbers: 13, 15 and 16, while the woody species studied mainly have 16, with no 13 recorded. Mangaly (1968) studied 8 herbaceous species in North America and reported 2n=26 for them except S. pseudo-china with 2n=30. Mangaly considered that a probably ancestral home of Smilax, both the herbaceous and woody, is in Southeast Asia and the eastern Himalayas, and speculated that the ancestral type of Sect. Coprosmanthus is possibly an Asian species, S. riparia. The karyotypes of the two herbaceous species in East Asia consist mostly of sm and m chromosomes, whereas those for the North American species are all of st chromosomes. Based on the general rule of karyotypic evolution, i.e. from symmetry to asymmetry, his speculation seems reasonable. Researches on sex-chromosomes of Smilax have been carried out since 1930 (Lindsay, 1930; Jensen, 1937; Nakajima, 1937; Mangaly, 1968), and they are generally considered to be the largest pair, but there is still no adequate evidence. The result of our observation on S. nipponica may confirm that the first pair of chromosomes of this species is XY type of sex-chromosomes. Chromosomes of the genus are small and medium-sized, varying between 1-6 μm, slightly larger in herbaceous species than in woody ones, larger in the karyotype of 2n=26 than in that of 2n=32. Based on karyotype constitution of the above 5 species, the karyotype in the genus is characterized by 4 pairs of L chromosomes and 2-5 pairs of M chromosomes, and mostly st and sm chromosomes, and by rather asymmetrical 3B type. The degree of symmetry in the above 5 species is from Sect. Coprosmanthus to Sect. Coilanthus, and herbaceous species towoody ones.     

7-Substituted pterins. A new class of mammalian pteridines

Three novel pteridines have been isolated from the urine of patients with a new variant of 6-(L-erythro-1',2'-dihydroxypropyl)-5,6,7,8-tetrahydropterin (tetrahydrobiopterin) deficiency, showing hyperphenylalaninemia. From the results of high performance liquid chromatography, oxidative degradation, and gas chromatography-electron impact mass spectrometry, their structures were identified as 7-(D-erythro-1',2',3'-trihydroxypropyl)-pterin (7-neopterin), 7-(L-erythro-1',2'-dihydroxypropyl)-pterin (7-biopterin), and 6-oxo-7-(L-erythro-1',2'-dihydroxypropyl)-pterin (6-oxo-7-biopterin). The ratio of biopterin to 7-biopterin in the patients' urines was 1:1, and after oral loading with tetrahydrobiopterin, 7-biopterin excretion rose parallel to biopterin. This finding suggests that 7-substituted pterins may be formed endogenously by a yet unknown isomerization reaction. The cause of hyperphenylalaninemia is still unclear. The activities of the enzymes involved in tetrahydrobiopterin biosynthesis and regeneration were found to be normal in the patients, and no effect of 7-biopterin on these enzymes was observed in vitro. However, compared with the normal cofactor, tetrahydrobiopterin, the Km values of tetrahydro-7-biopterin for phenylalanine hydroxylase and dihydropteridine reductase are 20 and 5 times higher, respectively.     

Studies of peptide antibiotics. XLI. Syntheses of 6-L-valine-tyrocidine A and 7-L-ornithine-tyrocidine A

Tyrocidine A (TA) is an antibiotic cyclic decapeptide with the sequence of cyclo (-L-Val1-L-Orn2-L-Leu3-D-Phe4-L-Pro5-L-Phe6-D-Phe7-L-Asn8-L-Gln9-L-Tyr10-). Gramicidin S (GS) regarded as a homolog of TA is also a cyclic decapeptide with the sequence of cyclo (-L-Val1-L-Orn2-L-Leu3-D-Phe4-L-Pro5-L-Val5-L-Orn7-L-Leu8-D-Phe9-L-Pro10-). GS shows higher antibacterial activity, whereas TA exhibits inhibitory activity on the biosynthesis of RNA. Two analogs of TA, [L-Val6]-TA (12a) and [L-Orn7]-TA (12b), were synthesized by the conventional method in order to study the interrelationships between the two related antibiotics TA and GS. Antibacterial activities of 12a and TA are nearly the same, but the activity of 12b is significantly lower. The optical rotatory dispersion spectra of 12a, 12b, and TA showed a trough at 233 nm region; the troughs of 12a and TA are nearly the same in depth, but the trough of 12b is shallower. Relationships between structure and activity of 12a and 12b compared with TA and GS were discussed.     

Engineering the exo-loop of Trichoderma reesei cellobiohydrolase, Cel7A. A comparison with Phanerochaete chrysosporium Cel7D

The exo-loop of Trichoderma reesei cellobiohydrolase Cel7A forms the roof of the active site tunnel at the catalytic centre. Mutants were designed to study the role of this loop in crystalline cellulose degradation. A hydrogen bond to substrate made by a tyrosine at the tip of the loop was removed by the Y247F mutation. The mobility of the loop was reduced by introducing a new disulphide bridge in the mutant D241C/D249C. The tip of the loop was deleted in mutant Delta(G245-Y252). No major structural disturbances were observed in the mutant enzymes, nor was the thermostability of the enzyme affected by the mutations.The Y247F mutation caused a slight k(cat) reduction on 4-nitrophenyl lactoside, but only a small effect on cellulose hydrolysis. Deletion of the tip of the loop increased both k(cat) and K(M) and gave reduced product inhibition. Increased activity was observed on amorphous cellulose, while only half the original activity remained on crystalline cellulose. Stabilisation of the exo-loop by the disulphide bridge enhanced the activity on both amorphous and crystalline cellulose. The ratio Glc(2)/(Glc(3)+Glc(1)) released from cellulose, which is indicative of processive action, was highest with Tr Cel7A wild-type enzyme and smallest with the deletion mutant on both substrates. Based on these data it seems that the exo-loop of Tr Cel7A has evolved to facilitate processive crystalline cellulose degradation, which does not require significant conformational changes of this loop.     

A kinetic study of thymine 7-hydroxylase from neurospora crassa.

The steady-state kinetics of thymine 7-hydroxylase (thymine, 2-oxoglutarate dioxygenase, EC 1.14.11.6) has been investigated. Initial velocity plots were all found to be linear and intersecting. Variation in concentration of two of the substrates, when the third substrate was at a constant high or low concentration, gave initial velocity plots that conform to an ordered sequential mechanism, where thymine is the second substrate to add. With 5-carboxyuracil, which is the end product in the sequential oxygenation of thymine, a competitive inhibition pattern was observed when 2-ketoglutarate was the variable substrate. When either thymine or oxygen was the variable substrate a noncompetitive inhibition pattern was obtained. When either 2-ketoglutarate or thymine was the variable substrate the inhibition patterns observed with bicarbonate were noncompetitive. With succinate noncompetitive inhibition patterns with hyperbolic intercept replots were obtained. These results are consistent with an ordered sequential kinetic mechanism, where 2-ketoglutarate is added first, followed by thymine and oxygen, and the products are released in the order: bicarbonate, succinate, and 5-hydroxymethyluracil. The order of the two last mentioned products, however, is changed in the presence of succinate.     

A radioimmunoassay for 7 alpha-hydroxy dehydroepiandrosterone in human plasma.

A radioimmunoassay for plasma 3 beta, 7 alpha-dihydroxy-5-androsten-17-one (7 alpha-hydroxy DHA) has been developed using anti-sera raised against 3 beta, 7 alpha-dihydroxy-5-androstene-17 beta-carboxyl-bovine serum albumin conjugate and [1, 2 (n) - 3H] 7 alpha-hydroxy DHA as the radioligand. Significant cross reactivity was found with 3 beta, 7 alpha-dihydroxy-5-pregnen-20-one (44%), 3 beta, 7 beta-dihydroxy-5-androsten-17-one (6%), 3 beta, 6 beta-dihydroxy-4-androsten-17-one (2.5%), 3 beta-hydroxy-5-androsten-17-one (DHA, 2%), 3 beta, 7 beta-dihydroxy-5-pregnen-20-one (2%) and 7 alpha-hydroxy-4-androstene-3, 20-dione (1%). 7 alpha-Hydroxy DHA was extracted from plasma and separated from cross-reacting factors using alumina micro-columns. The separation of bound and free steroid was achieved using dextran-coated charcoal. The concentration of 7 alpha-hydroxy DHA in the plasma of breast cancer patients was significantly lower than the concentrations in the plasma of normal women, hospitalized women suffering from non-endocrine diseases and patients with benign breast disease. The decrease in the concentration of 7 alpha-hydroxy DHA in the plasma of pregnant women was not significant.