Introduction of rice chitinase gene into wheat via low energy Ar+ beam implantation

A chitinase gene (RCH8) in plasmid vector pCAMBIA1308 was delivered into 3 wheat cultivars (Yangmai 158, Wan 9210, Wanmai 32) by low energy Ar⁺ beam-mediated method. Preliminary calli from treated mature embryos were first selected on hygromycin (Hm, 20 or 30 mg/L) containing medium. After the resistant calli formed, they were transferred to the regeneration medium with 10 or 20 mg/L Hm. All the three wheat varieties obtained transgenic plants. PCR and PCR-Southern assays showed that most plants regenerated from the resistant calli were positive transgenic plants. Southern blot of the positive green plants confirmed stable integration of alien DNA into wheat genome. The plant transformation frequencies varied with the variety and ion dose implanted. Wanmai 32 possessed the highest transformation frequency, reaching 3.8% at a suitable implantation dose. The transformation frequency of Yangmai 158 and Wan 9210 varied from 0.5% to 2.5% and from 0.5% to 1.4%, respectively. Progeny test for resistance to wheat scab showed that the leaf extract of R₁ generation inhibited the growth of wheat scab strain R₀ and F₁₅.     

The pathogenic site of the C-toxin derived from Bipolaris maydis race C in maize(Zea mays)

Bipolaris maydis race C strain 523 (C523) induces severer leaf blight on cytoplasmic male sterility (CMS)-C maize than on normal (N) maize. Previously, a pathotoxin isolated from C523 (C-toxin) was shown to be responsible for the disease. To understand the basis of the differential responses between CMS-C and N maizes to this fungus, protein synthesisin vitro by mitochondria from N and CMS-C cytoplasms was monitored after their incubation in a solution containing the toxin (0.3%). Similar protein products were detected between the two alloplasmic lines, indicating that the toxin does not directly act on the mitochondrial membrane, nor inhibits the expression of mitochondrial genes. To further locate the action site of the toxin, intact leaves from both N and several subtypes of CMS-C lines were treated by 0.3% toxin. Analysis of electrolyte leakage of leaf cells showed that the leakage rates were similar to one another among the alloplasmic maize lines. In contrast, at a lower concentration of the toxin (0.05%), the leaf cells from CMS-C line were more susceptible to the toxin than those of the other lines. All these results indicate that the target of the toxin action appears to be the cellular-membrane rather than mitochondria, suggesting that the variable susceptibilities toB. maydis between the alloplasmic maize lines might be related to a difference in their cellular-membranes.     

Studies on the utilization potentiality of the nucleo-cytoplasmic hybrid in wheat

A series of comparative studies was carried out on the genetic effects of 25 alien cytoplasms of wheat on the growth potential, heading stage, fertility, resistance against diseases, important agronomic traits and its heterosis of 125 nuclei-cytoplasmic hybrids of wheat. The results indicated that there were clearly effects of alien cytoplasms on some characteristics, but the nucleus still exerted main effect on other characteristics. The effect of interactions between nucleus and cytoplasm was comparative obvious in some combination. Consequently, when we utilize the effects of alien cytoplasms, we should pay full attention to the facts such as the characteristic to be improved, the effects of cytoplasm, nucleus, the nucleus-cytoplasm interactions on that characteristics. From the preliminary studies, we believed that the cytoplasmic types of M⁰, S¹, S^v, D², D and B, and the nucleo-cytoplasmic hybrids of (Ae. sharonensis)-B174, (Ae. squarrosa)-352-35 (Ae. cylindrica)-352-35, (Ae. cylindria)-E EN-1, (Ae. cylindrica)-NPFP, and (Ae. speltoides)-352-35 would have some utilization potentiality in cultivar improvement. Foundation item: supported by the Foundation of National 8th Five-year-plan of China Biography: CHEN Jun-ying, (1964-), male, lecturer, and now worked in Henan Agricultural University, Zhengzhou 450002, China     

Restorable variation of cytoplasmic type of male sterile line in rice byin vitro culture

Rice male sterile (MS) lines, IR66707A and IR69700A, which possess the cytoplasm of Oryza perennis and O. glumaepatula respectively, belong to the cytoplasmic type. Their sterility could be maintained but not be restored. By somatic cell culture of these two MS lines, 47 somaclones with 465 R₁ plants were obtained. All of the 465 R₁ plants were sterile in the spring season in Guangzhou. According to the expression of the R₁ plants and the level of similarity to their donor parents, they could be divided into three types. The plants of type Ⅰ were male sterile. The sterility of some somaclones of this type could be restored by the test crossing varieties or alternated to fertile by changes of some environmental conditions. The hybrid F₁ of test cross from the MS somaclones in type Ⅰ was fertile while the hybrid F₁ from the donor MS lines was still sterile. The R₁ plants of type Ⅱ were similar to the donor parents and also male sterile. The hybrid F₁ from all of the plants of type Ⅱ crossed to test variety were still sterile, so they did not possess restorability. For the somaclone of type Ⅲ, all of R₁ plants were sterile in both male and female organs. No seed was set in both conditions of self and cross pollination. The fact that the restorable variants obtained in the cytoplasmic type of MS lines of rice by in vitro culture reported here should be the first sample in somaclonal variation in plant kingdom.     

Relationships between D1 protein, xanthophyll cycle and photodamage-resistant capacity in rice (Orysa sativa L.)

Relationships between D1 protein, xanthophyll cycle and subspecific difference of photodamage-resistant capacity have been studied inO. japonica rice varieties 02428 and 029 (photoinhibition-tolerance) andO. indica rice varieties 3037 and Palghar (photoinhibition-sensitivity) and their reciprocal cross F₁ hybrids after photoinhibitory treatment. It was shown that PS II photochemical efficiency (F _v /F _m) decreased, and xanthophyll cycle from violaxanthin (V), via anaxanthin (A), to zeaxanthin (Z) was enhanced and non-photochemical quenching (qN) increased accordingly in SM-pretreated leaves of rice when the synthesis of D1 protein was inhibited, and that there was a decrease inqN and, as a result, more loss of D1 protein and a big decrease inF _v/F _m in DTT-pretreated leaves when xanthophyll cycle was inhibited.O. japonica subspecies had a higher maintaining capacity of D1 protein and a decrease ofF _v/F _m in a more narrow range, and exhibited more resistance against photodamage, as compared withO. indica subspecies. The above physiological indexes in reciprocal cross F₁ hybrids, though between the values of their parents, were closer to maternal lines than to paternal lines. Experimental results support the concept that the turnover capacity for D1 protein is an important physiological basis of photoinhibition-tolerance, and will provide the physiological basis for selection of the photoinhibition-tolerant parents and develop a new approach to breed hybrids with high photosynthetic efficiency.     

Molecular cytogenetic analysis of Triticum aestivum-Leymus racemosus reciprocal chromosomal translocation T7DS·5LrL/T5LrS·7DL

In order to induce chromosome translocation between wheat chromosomes and chromosome 5Lr of Leymus racemosus, the mi- crosporocytes during meiosis of T. aestivum-L. racemosus disomic addition line DA5Lr were irradiated by 60Co γ-rays 800 R (100 R/min). Before flowering, the treated spikes were emasculated and bagged. After 2-3 d, the emasculated flowerets were pollinated using pollens from T. aestivum cv. Chinese Spring. One plant with two translocation chromosomes involved in both the long and short arm of...     

K型小麦雄性不育系的细胞质效应

Ｋ质对杂种ＦＱ开花期旗叶的净光合速率和呼吸速率无不良影响,在与一定核作用下,能够提高净光合速率。     

Development of Triticum aestivum-Leymus racemosus ditelosomic substitution line 7Lr#1S（7A） with resistance to wheat scab and its meiotic behavior analysis

Leymus racemosus is highly resistant to wheat scab （Fusarum head bright）. The transfer of scab resistant gene from L. racemosus to Triticum aestivum is of great significance for broadening the base of wheat resistance. In the present study, the pollen of T. aestivum-L, racemosus monosomic addition line with scab resistance was treated by irradiation with 1200 R ^60Co-γ-rays prior to pollinating to emasculated wheat cv. Mianyang 85-45. Nine plants with a telocentric chromosome 7Lr#1S were observed in M1, and one ditelosomic substitution line 7Lr#1S was selected from selfcrossing progenies and confirmed by chromosome C-banding and GISH. Furthermore, a co-dominant EST-SSR marker CINAU 31 was employed to identify this substitution line. A pair of chromosome 7A of common wheat were found to be replaced by a pair of telocentric chromosome 7Lr#1S, and further investigation showed that chromosome configuration of the substitution line at MI of PMCs after GISH was 17.50Ⅱ^w ＋ 2.19Ⅱ^w ＋ 0.42Ⅱ^7Lr#1S ＋ 1.08 Ⅰ^7Lr#1S ＋ 0.69 Ⅰ^w. Two telocentric chromosomes paired as a bivalent in 59.7% of PMCs. Abnormal chromosome behaviors of telocentric chromosomes were observed in part of PMCs at anaphase I and telophase I, including the moving of two telocentric chromosomes to the same pole, lagging and earlier separation of their sister chromatid. All these abnormal behaviors can be grouped into three distinct types of tetrads according to different numbers of 7Lr#1S in their daughter cells and various micronucleus in some tetrads. However, due to the high transmission frequency of the female and male gametes with a 7Lr#1S, 84% of the selfcrossing progeny plants had ditelosomic substitution. The substitution line showed high resistance to wheat scab in a successive two-year test both in the greenhouse and field; hence, the line will be particularly valuable for alien gene mapping, small fragment translocation induction and telosomic cytological behavior analysis.     

渗透胁迫对小麦愈伤组织GST活性的影响

比较了渗透胁迫下抗旱性不同的小麦愈伤组织GST活性的变化。结果表明 ,水分胁迫初期2种愈伤组织GST活性均明显高于对照 ,重度胁迫的高于轻度胁迫 ;但在轻度胁迫下的后期小偃22细胞GST活性明显高于对照或与对照无明显差别 ,而郑引1号GST活性明显低于对照 ;说明GST可能与细胞对渗透胁迫的适应有关     

Tagging major quantitative trait loci for sheath blight resistance in a rice variety, Jasmine 85

This study was conducted with a clonal F₂ population of rice from a cross between Jasmine 85, a resistant variety, and Lemont, a susceptible cultivar. The rice plants belonging to each F₂ clone were divided into two plots, which were put in two replicates, respectively. Clonal parents were tested as controls. The plants were inoculated by short toothpicks incubated with RH-9, a virulent isolate of the pathogenic fungus,Rhizoctonia solani, which causes rice sheath blight. The extreme resistant and susceptible clonal lines were selected for construction of resistant and susceptible DNA pools, respectively. A total of 94 polymorphic markers evenly distributed on 12 rice chromosomes were used for bulked segregant analysis, three positive ones were found polymorphic between the two DNA pools, and three major QTLs for sheath blight resistance, Rh-2, Rh-3 and Rh-7, were identified. The three major QTLs were located on chromosomes 2, 3 and 7, and could explain 14.4%, 26.1% and 22.2% of the phenotypic variation.     

Molecular mapping of a novel yellow rust resistance gene of wheat using microsatellite markers

Identification and genetic analysis of yellow rust resistance have suggested that wheat line R55 carries single dominant gene conferring yellow rust resistance. The bulked segregant analysis (BSA) for an F₂ population using microsatellite marker technique has indicated that the yellow rust resistance gene is located on the short arm of chromosome 1B, tightly linked to the microsatellite markers WMS11-193 bp and WMS18-184 bp, the linkage distance between the markers and the gene is 1.9 cM. This gene has been formally namedYr26. It is inferred from the pedigree, resistance and gene locus analysis that theYr26 has been transferred fromTriticum turgidum L. and is different from the other known yellow rust resistance genes.     

Breeding and molecular cytogenetic identification of wheat-Thinopyrum intermedium addition lines resistant to powdery mildew

Wheat-related species Th. intermedium was used to cross with common wheat Yannong 15. In the self progenies of the hybrid, two addition lines, Ⅱ-1-7-1 and Ⅱ-3-3-2, stable in cytology, were developed by cytology and powdery mildew resistance identification. Their chromosome number were 2n = 44 and formed 22 bivalents at PMC MI. In F₁ of the two addition lines crossing with Yannong 15, there appeared about one univalent at PMC MI, respectively. Resistance identification in greenhouse and field using the No. 15 and mixed strains of E. gramnis f. sp. tritici showed that they were immune to powdery mildew. Chromosome number and resistance identification using the F² single plants of the addition line crossing with Yannong 15 indicated that the resistant gene was located on the alien chromosomes. In situ hybridization using St and E genomic DNA as probe showed that the added chromosome in the two addition lines probably came from the E genome of Th. intermedium, which indicated that a pair of E genome chromosomes carried a new resistant gene to powdery mildew.     

Two-dimensional crystallization and preliminary electron crystallographic result of partially purified F<Subscript>0</Subscript> from porcine mitochondria

After removal of cytoplasmic sector F₁ from submitochondrial particles of F₀F₁-ATP synthase complex with guanidine hydrochloride, the transmembrane sector F₀ was specifically extracted from the stripped membranes in the presence of detergent CHAPS and partially purified. Two-dimensional crystals were produced by the reconstitution of the partially purified F₀ into asolectin and microdialysis. The obtained crystals are able to diffract to 2 nm. The projection map of the negatively stained crystal shows that the crystal has p42₁2 symmetry, lattice constant, a=b=14.4 nm. A unit cell contains four F₀ molecules.     

Gene expression profiling related to powdery mildew resistance in wheat with the method of suppression subtractive hybridization

“Bainong 3217 × Mardler” BC₅F₄ wheat line at the initial stage of inoculation with powdery mildew pathogen (Erysiphe graminis DC) was used to construct a suppression subtractive hybridization (SSH) cDNA library. Totally 760 ESTs were obtained through sequencing. Similarity analysis of ESTs based on BLASTn and BLASTx with the sequences in GenBank, in combination with macroarray differential screening, revealed that 199 ESTs of 65 kinds were known to be functionally disease resistance related. Based on the gene expression profiling in the present study, it is postulated that salicylic acid (SA) and MAP-related signal transduction pathways were involved in powdery mildew resistance in wheat. System acquired resistance genes were predominant in terms of kinds and quantity. With the initiation of cell defense reaction, the genes conferring anti-oxidation substances were largely expressed and thus cell protection mechanism was activated. Much evidence revealed that phenylpropanes metabolic pathway was involved in phytoalexin synthesis in wheat powdery mildew resistance. Genes conferring some enzymes of structural modification of cell walls and proteinase inhibitors inhibiting pathogen growth were also detected. The genes controlling a few proteinases (mainly cysteine proteinase) had a considerable redundancy of expression.     

Inheritance of resistance toHelicoverpa armigera of 3 kinds of transgenicBt strains available in upland cotton in China

There are 3 kinds of transgenicBt strains, Shanxi 94-24, Zhongxin 94, and R19, in upland cotton in China. Their transgenicBt insect-resistance cultivars or hybrids have been developed and grown by farmers. Genetic studies indicate that the resistance of the 3 transgenicBt cotton strains toHelicoverpa armigera is controlled by one pair of non-allelic dominant genes. Linkage relationship between the resistant genes of R19 and Shanxi 94-24 transgenicBt strains shows that they may be inserted in the same chromosome. F₁ hybrids crossed among the 3 strains show that high levels of protection from feeding damage are the same as that of their parents. Therefore, there is no co-suppression phenomenon in many transgenic plants. The results presented here afford a fundamental reliance in developing transgenicBt insect-resistant cultivars and exploiting the heterosis of hybrids in upland cotton.     

Identification of an AFLP marker linked to the stripe rust resistance geneYr10 in wheat

AFLP analysis of near-isogenic lines of the stripe rust resistance gene Yr10 was carried out with 6 PstⅠ- primers and 10 TaqⅠ-primers with the donor parent of Yr10 gene as the check. A total of about 4200 distinguishable bands were amplified, of which 5 were stable. The genetic linkage of the 5 polymorphic DNA fragments with the target gene were tested preliminarily on a segregating F₂ population derived from a cross between the gene donor parent “Moro” and susceptible cultivar “Mingxian 169”. The DNA fragment PT0502 was found closely linked to the Yr10 gene and cloned and sequenced. Based on the sequence specific primers for PCR were designed and synthesized. Genetic linkage analysis with 195 segregating F₂ plants indicated that the genetic distance was 0.5 cM between the main product SC₂₀₀ fragment produced by PCR with the primers and the Yr10 gene. The primers can be used to detect the Yr10 gene quickly, effectively and exactly.     

Breeding of T. aestivum-Ag. intermedium translocation lines with T. timopheevii cytoplasm and characterization by GISH

A male-sterileT. aestivum-Ag. intermedium partial amphiploid with cytoplasm ofT. timopheevii as a female parent was crossed to common wheat. The hybrid was backcrossed to the male parent several times continually and setf-crossed at last. Two stable lines with common wheat phenotype, H96269-2 and H96278, have been obtained. The chromosome numbers of the two lines are all2n = 42 in somatic cetls. By inoculation test, the two lines show a high levet of resistance to yetlow rust. Through genomicin situ hybridization (GISH) withAg. intermedium total genomic DNA as a probe, it is demonstrated that the two stable lines are all small segmental translocation lines, and the translocated chromosome segments fromAg. intermedium are located on the short arm terminals of wheat chromosomes. Genetics analysis suggests that the yetlow rust resistance gene(s) are probably located on the translocated chromosome segments ofAg. intermedium.     

Genetic transformation of calli from maize and regeneration of herbicide-resistant plantlets

The herbicide-resistant geneals ofArabidopsis thaliana has been transferred into embryonic calli of maize by microprojectile bombardment. We have obtained chlorsulfuron-resistant calli and regenerated plantlets through selection by herbicide chlorsulfuron. The results of PCR analysis and Southern blotting indicate that geneals has been transferred into some plantlets. The test of spraying chlorsulfuron indicated that the transgenic plantlets had favorable herbicide-resistant trait. The purpose of the research was to obtain chlorsulfuron-resistant transgenic maize and hope that this kind of high efficient herbicide could be widely used in rotation soil of wheat and maize. In addition, through spraying herbicide, we could eliminate the hybrid plants and thereby increase the purity of F1 seeds.     

Fine mapping of the red plant gene R1 in upland cotton(Gossypium hirsutum)

Sub 16 is a substitution line with G. hirsutum cv. TM-1 genetic background except that the 16th chromosome (Chr. 16) is replaced by the corresponding homozygous chromosome of G. barbadense cv. 3-79, and T586 is a G. hirsutum multiple gene marker line with 8 dominant mutation genes. The R ₁ gene for anthocyanin pigmentation was tagged in Chr. 16 in T586. The objective of this research was to screen SSR markers tightly linked with R ₁ by using the F₂ segregating population containing 1259 plants derived from the cross of Sub 16 and T586 and the backbone genetic linkage map from G. hirsutum×G. barbadense BC₁ newly updated by our laboratory. Genetic analysis suggested that the segregation ratio of red plants in the F₂ population fit Mendelian 1:2:1 inheritance, confirming that the red plant trait was controlled by an incomplete dominance gene. Preliminary mapping of R ₁ was conducted using 237 randomLy selected F₂ individuals and JoinMap v3.0 software. Then, a fine map of R1 was constructed using the F₂ segregating population containing 1259 plants, and R ₁ was located between NAU4956 and NAU6752, with only 0.49 cM to the nearest maker loci (NAU6752). These results provided a foundation for map-based cloning of R ₁ and further development of cotton cultivars with red fibers by transgenic technology. Supported by National Natural Science Foundation of China (Grant No. 30730067) and Programme of Introducing Talents of Discipline to Universities (Grant No. B08025)     

Mg2+-induced LHC II aggregation in thylakoid membrane

Mg²⁺-induced changes in 77K fluorescence spectra of wheat thylakoid membranes were analyzed by a Gaussian deconvolution program. All spectra were fitted well with 7 Gaussian sub-bands. The sub-band areas for the LHC H macroaggregate (F₆₉₉) and PS II (F₆₈₅ and F₆₉₅) were increased and those for the LHC II trimer (F₆₈₁) and PS I (F₇₂₉ and F₇₄₀) were decreased by Mg²⁺. Moreover, it was found that the sub-band area ratio for F₆₉₉ over F₆₈₁ was dramatically enhanced by Mg²⁺ by 1.72 times. It was concluded that the LHC II trimer in thylakoid membranes could be aggregated and transformed into its macroaggregate by Mg²⁺ on a large scale. The possible implication of the Mg²⁺-induced LHC II aggregation was also discussed.