水稻苗期耐低钾品种筛选及相关性状的QTL定位

对159份遗传差异较大的水稻品种进行耐低钾筛选,根据综合指数筛选出18份耐低钾和10份低钾敏感的水稻品种。利用筛选得到的耐低钾水稻品种台农67和低钾敏感品种制西构建重组自交系,对耐低钾相关性状进行QTL分析和定位,共检测到27个耐低钾相关性状的QTL,LOD值为2.52～9.23,可解释表型变异为2.22%～7.25%,其中贡献率最高的QTL是qKC-2,该QTL定位在水稻2号染色体上,可能是1个控制水稻钾含量的未知基因,可进一步通过构建次级群体对该基因进行精细定位和克隆。本研究中设计的Indel标记可快速筛选水稻育种材料中的耐低钾基因型,缩短育种时间,而且重组自交系中可以选出农艺性状好又耐低钾的株系直接应用于育种。     

全球水稻分子育种核心种质资源耐低钾品种的苗期筛选

以全球水稻分子育种计划提供的117份核心种质资源为供试材料,进行苗期水培试验,比较两种不同K 浓度处理下的苗期根茎性状表现.结果表明:两种处理条件下,品种间存在显著差异;以发根数、总根长、苗高、地下部干重、地上部干重等5个性状相对指数的平均值构成品种的综合指数,作为筛选耐低钾品种的评判标准,结合稻苗生长情况,从117份核心种质资源中筛选出9个耐低钾品种和32个较耐低钾品种.     

营养胁迫下东乡野生稻生物学特性鉴定初报

以东乡野生稻中东塘、东塘西侧2个居群和耐低磷品种大粒稻、低磷敏感品种新三百粒为材料,采用砂培法和土培法,鉴定营养胁迫下东乡野生稻的生物学特性。砂培试验表明,在低氮、低磷、低钾胁迫下,东塘、东塘西侧地上部干重、生物学产量降低;根冠比显著增大,其中低氮胁迫下表现尤为明显。土培试验表明,低氮胁迫下株高和分蘖动态变化与无肥基本一致;各胁迫下,大粒稻和新三百粒分蘖后期有分蘖数下降阶段,东塘和东塘西侧的分蘖数基本保持不变;低磷胁迫下,东塘叶重、根重、茎鞘重和东塘西侧剑叶长、叶重、茎重、总产量性状与全肥处理相比差异显著。其他性状与全肥处理无显著差异。低钾胁迫下,东塘除叶重、根重外其他11个性状与全肥处理无显著差异;东塘西侧除着粒密度、穗重外其他11个性状与全肥处理无显著差异。初步表明,东塘和东塘西侧2个居群无耐低氯特性,具有耐低磷、低钾特性,且不同居群时低磷、低钾的忍耐能力有一定差异。     

拟南芥钾转运突变体atkup12鉴定及非生物胁迫的敏感性检测

[目的]鉴定获得拟南芥HAK/KUP/KT高亲和钾离子转运突变体atkup12,通过检测种萌期atkup12突变体在低钾、盐及氧化胁迫下的生长指标以初步明确拟南芥AtKUP12基因是否参与植物对非生物胁迫的响应。[方法]以拟南芥atkup12突变体基因组和总RNA反转后的cDNA为模板,通过PCR扩增确定AtKUP12基因T-DNA插入失活的纯合突变体。将野生型和atkup12突变体点种于0.5μmol/L低钾、不同NaCl浓度和1μmol/L甲基紫精的胁迫培养基,测定并比较突变体与野生型拟南芥在根长、种子萌发率及子叶绿化率间的差异。[结果]利用双引物法,结合反转录PCR,在DNA及RNA水平鉴定获得了AtKUP12基因的纯合突变体。低钾胁迫下,atkup12突变体种苗的根长比较野生型拟南芥短40%左右,较野生型受到了显著抑制;不同NaCl浓度胁迫下,atkup12突变体种子的萌发率较野生型显著降低;1μmol/L甲基紫精显著的抑制了突变体种苗的子叶绿化率。[结论]成功鉴定获得了AtKUP12基因T-DNA插入失活的纯合突变体,且AtKUP12基因的缺失会增加植物对盐、低钾及氧化等非生物胁迫的敏感性。     

水稻直链淀粉和蛋白质突变系筛选及其与主要农艺性状的关联分析

研究利用重离子辐照杂交籼稻9311创建农艺性状突变体库,使用化学方法筛选直链淀粉、蛋白质突变材料,并分析籽粒品质性状与农艺性状之间的相关性,为后续筛选直链淀粉、蛋白质突变体工作奠定基础。结果显示:在169份直链淀粉、蛋白质突变体中,直链淀粉含量变幅范围是7.64%~32.37%,其中高含量直链淀粉突变体材料有11份,低含量直链淀粉突变体材料有5份;蛋白质含量变幅范围是7.05%~13.79%,其中高蛋白突变体有34份,低蛋白突变体有3份。从169份突变体中筛选出直链淀粉、蛋白质含量都有梯度差异的突变体材料,筛选出材料的农艺性状分析结果表明:在直链淀粉、蛋白质突变体材料的农艺性状中,每穗实粒数、有效穗数、结实率和株高这四个农艺性状与籽粒品质性状间有关联,变异系数分别为37.53%、30.72%、24.70%、15.38%。相关性结果表明:直链淀粉含量和蛋白质含量呈负相关,直链淀粉含量与农艺性状呈正相关,蛋白质与农艺性状呈负相关,其中蛋白质含量和每穗实粒数、结实率呈极显著负相关,相关系数分别为-0.504、-0.592。     

棉花抗枯、黄萎病品种苗期耐低钾种质筛选研究

本研究采用蛭石栽培和营养液浇灌的方法,确立棉花品种苗期耐低钾筛选指标,利用这些指标,评价我国88个抗枯、黄萎病棉花品种资源苗期耐低钾能力,并筛选钾素利用率高的优异种质。结果表明,株高、地上部干物重、地下部干物重、总干物重、钾利用率、叶绿素含量在不同钾浓度处理和不同品种之间均存在显著差异,可以作为棉花苗期耐低钾能力的评价指标,而叶面积在不同品种和不同钾浓度处理之间不存在差异。利用筛选出的指标对88个品种进行耐低钾评价,结果供试品种主要被分为耐低钾基因型、耐低钾中间类型和非耐低钾基因型3类,分别包括21个、58个和6个品种。     

棉花抗枯、黄萎病品种苗期耐低钾种质筛选研究

本研究采用蛭石栽培和营养液浇灌的方法,确立棉花品种苗期耐低钾筛选指标,利用这些指标,评价我国88个抗枯、黄萎病棉花品种资源苗期耐低钾能力,并筛选钾素利用率高的优异种质。结果表明,株高、地上部干物重、地下部干物重、总干物重、钾利用率、叶绿素含量在不同钾浓度处理和不同品种之间均存在显著差异,可以作为棉花苗期耐低钾能力的评价指标,而叶面积在不同品种和不同钾浓度处理之间不存在差异。利用筛选出的指标对88个品种进行耐低钾评价,结果供试品种主要被分为耐低钾基因型、耐低钾中间类型和非耐低钾基因型三类,分别包括21个、58个和6个品种。     

常夏石竹耐盐突变体渗透调节的研究

在离体培养条件下利用γ-射线作诱变剂获得耐0.5%、0.7%、1.0%NaCl的突变系,通过对稳定突变系植株叶片渗透剂含量及对渗透势贡献大小的测定表明耐盐突变体叶中K+、游离氨基酸、Na+、脯氨酸的含量高于对照,其中脯氨酸和Na+积累最明显。而叶片中可溶性糖的含量、K+/Na+低于对照。Na+对突变体植株叶片渗透势贡献最大,是最主要的渗透调节剂之一。耐盐突变体植株内存在渗透物质的再分配,叶内有吸钾排钠现象。     

不同绿豆突变体主要农艺性状的多元遗传分析

对12个绿豆突变体的10个主要农艺性状进行了相关分析、主成份分析及聚类分析.分析结果表明：10个主要农艺性状的变异系数为16.01%（单株荚数）～3.64%（荚宽）.相关分析表明单株荚数与单株产量呈极显著的正相关;百粒重与单株产量呈显著的负相关;生育期与单株产量呈极显著的负相关.主成分分析结果表明前4个主成分（产量构成因子、单荚粒数因子、株型因子和荚宽因子）对变异的贡献率达87.45%;聚类分析后把12个绿豆突变体分为5类,各类之间单株产量差异明显.在绿豆突变体的选择中只有把不同性状综合考虑进去,才能够真正选择出性状优良的突变体.     

空间诱变水稻大粒型突变体的遗传育种研究

对粳稻农垦58经空间诱变产生的大粒型突变体进行大粒型性状的遗传分析和育种应用研究。结果表明,大籽型突变体的籽粒大小（以籽粒体积表示）表现为受多基因控制的数量性状。大粒型突变体谷粒细长,具馨香味,外观品质优,对粳稻的亲和性好,是一个罕见的优质米资源。以大粒型突变体为供体,高产、外观米质差的晚粳推广品种为受体,采用不饱和回交结合分离世代糙米外观品质鉴定的方法,将大粒型突变体的优质性状导入晚粳背景,选育出外观米质优的晚粳新品系。对采用不饱和回方法改良数量性状进行了讨论。     

Isolation and Genetic Analysis of Arabidopsis Mutants with Low-K+ Tolerance

Ethyl methane-sulfonate (EMS)-mutagenized Arabidopsis M2 populations were screened in low-K+ medium using the root-bending assay. Forty-two putative low-K+-tolerant ( lkt ) mutants were selected from 150?000 tested M2 seedlings, and two of these mutants maintained their low-K+-tolerant phenotype in their M3 generations, respectively. Genetic analysis showed that either one of these two mutants has a monogenic recessive mutation in a nuclear gene, and that the two mutations in two independent mutants are allelic to each other.     

Mutants of Ralstonia (Pseudomonas) solanacearum sensitive to antimicrobial peptides are altered in their lipopolysaccharide structure and are avirulent in tobacco.

Ralstonia solanacearum K60 was mutagenized with the transposon Tn5, and two mutants, M2 and M88, were isolated. Both mutants were selected based on their increased sensitivity to thionins, and they had the Tn5 insertion in the same gene, 34 bp apart. Sequence analysis of the interrupted gene showed clear homology with the rfaF gene from Escherichia coli and Salmonella typhimurium (66% similarity), which encodes a heptosyltransferase involved in the synthesis of the lipopolysaccharide (LPS) core. Mutants M2 and M88 had an altered LPS electrophoretic pattern, consistent with synthesis of incomplete LPS cores. For these reasons, the R. solanacearum gene was designated rfaF. The mutants were also sensitive to purified lipid transfer proteins (LTPs) and to an LTP-enriched, cell wall extract from tobacco leaves. Mutants M2 and M88 died rapidly in planta and failed to produce necrosis when infiltrated in tobacco leaves or to cause wilting when injected in tobacco stems. Complemented strains M2* and M88* were respectively obtained from mutants M2 and M88 by transformation with a DNA fragment harboring gene rfaF. They had a different degree of wild-type reconstituted phenotype. Both strains retained the rough phenotype of the mutants, and their LPS electrophoretic patterns were intermediate between those of the wild type and those of the mutants.     

Studies on three mutagen-sensitive mutants of mouse L5178Y cells. II. Complementation analyses between two methyl methanesulfonate-sensitive mutants and between two 4-nitroquinoline-1-oxide-sensitive mutants

Three mutagen-sensitive mutants, MS-1, M10 and Q31, were isolated from mouse L5178Y cells. MS-1 cells are sensitive to methyl methanesulfonate (MMS), M10 cells are cross-sensitive to X-rays, MMS and 4-nitroquinoline-1-oxide (4NQO); and Q31 cells are cross-sensitive to UV and 4NQO. MMS-, X-ray- and UV-sensitive markers in these mutants behaved recessively in hybrids between pairs of these mutants as in hybrids between L5178Y and these mutants as reported before (Shiomi et al., 1982b). Complementation analyses were carried out by forming hybrids between two MMS-sensitive mutants (MS-1 and M10) and between two 4NQO-sensitive mutants (M10 and Q31). MMS and 4NQO survivals were measured in these hybrid cells. MS-1 and M10 were found to belong to different complementation groups for MMS-sensitive phenotypes. The hybrid clones between M10 and Q31 were as sensitive to 4NQO as each of the mutants, indicating codominance of 4NQO sensitivity in these mutants. The hybrids constructed with L5178Y and three mutants were stable as to their chromosome constitution for 100 days of cultivation without selective pressure. From the segregation studies on these hybrids, it is concluded that neither the X-ray-sensitive mutation in M10 nor the UV-sensitive mutation in Q31 is located on the X chromosome.     

Cucumber mosaic virus-plant interactions: identification of 3a protein sequences affecting infectivity, cell-to-cell movement, and long-distance movement

Mutants of the Cucumber mosaic virus (CMV) movement protein (MP) were generated and analyzed for their effects on virus movement and pathogenicity in vivo. Similar to the wild-type MP, mutants M1, M2, and M3, promoted virus movement in eight plant species. Mutant M3 showed some differences in pathogenicity in one host species. Mutant M8 showed some host-specific alterations in movement in two hypersensitive hosts of CMV. Mutant M9 showed altered pathogenicity on three hosts and was temperature sensitive for long-distance movement, demonstrating that cell-to-cell and long-distance movement are distinct movement functions for CMV. Four mutants (M4, M5, M6, and M7) were debilitated from movement in all hosts tested. Mutants M4, M5, and M6 could be complemented in trans by the wild-type MP expressed transgenically, although not by each other or by mutant M9 (at the restrictive temperature). Mutant M7 showed an inability to be complemented in trans. From these mutants, different aspects of the CMV movement process could be defined and specific roles for particular sequence domains assigned. The broader implications of these functions are discussed.     

Identification of a glucokinase that generates a major glucose phosphorylation activity in the cyanobacterium Synechocystis sp. PCC 6803

In silico analysis of genome of the cyanobacterium Synechocystis sp. PCC 6803 identified two genes, slr0329 and sll0593, that might participate in glucose (Glc) phosphorylation (www.kazusa.or.jp/cyano). In order to determine the functions of these two genes, we generated deletion mutants, and analyzed their phenotypes and enzymatic activities. In the presence of 10 mM Glc, wild-type (WT) and slr0329 defective strain (M1) grew fast with increased respiratory activity and NADPH production, whereas the sll0593 deletion mutant (M2) failed to show any of the Glc responses. WT and M1 were not significantly different in their glucokinase activity, but M2 had 90% less activity. Therefore, we propose that Sll0593 plays a major role in the phosphorylation of glucose in Synechocystis cells.     

Phosphatase of Chlamydomonas reinhardi: biochemical and cytochemical approach with specific mutants.

The unicellular alga Chlamydomonas reinhardi produces two constitutive acid phosphatases and three depressible phosphatases (a neutral and two alkaline ones) that can utilize napthyl phosphate as a substrate. Specific mutants depressible phosphatase were used to investigate biochemical properties and the cytochemical localization of these enzymes. The two constitutive phosphatases show similar pH optima (about 5.0) and Km values (2 x 10(-3) to 3.3 x 10(-3) M) but differ in their heat sensitivity and affinity for glycerophosphate.     

Thermodynamic analysis of helix-engineered forms of the activation domain of human procarboxypeptidase A2.

Thermodynamic characterization of the activation domain of human procarboxypeptidase A2, ADA2h, and its helix-engineered mutants was carried out by differential scanning calorimetry. The mutants were engineered by changing residues in the exposed face of the two alpha helices in order to increase their stability. At neutral and alkaline pH the three mutants, alpha-helix 1 (M1), alpha-helix 2 (M2) and alpha-helix 1 and alpha-helix 2 (DM), were more stable than the wild-type domain, in the order DM, M2, M1 and wild-type. Under these conditions the CD and NMR spectra of all the variants are very similar, indicating that this increase in stability is not the result of gross structural changes. Calorimetric analysis shows that the stabilizing effect of mutating the water-exposed surfaces of the helices seems to be mainly entropic, because the mutations do not change the enthalpy or the increase in heat capacity of denaturation. The unfolding behavior of all variants changes under acidic conditions: whereas wild-type and M1 have a strong tendency to aggregate, giving rise to a beta conformation upon unfolding, M2 and DM unfold reversibly, M2 being more stable than DM. CD and NMR experiments at pH 3.0 suggest that a region involving residues of the second and third beta strands as well as part of alpha-helix 1 changes its conformation. It seems that the enhanced stability of the altered conformation of M2 and DM reduces the aggregation tendency of ADA2h at acidic pH.     

Revertants and Secondary arom-2 Mutants Induced in Non-Complementing Mutants in the arom Gene Cluster of Neurospora Crassa

Extensive genetical and biochemical studies have been performed with revertants and secondary arom-2 mutants induced in two different primary non-complementing mutants which map within the arom gene cluster of Neurospora crassa. These studies indicate that mutant M54 but not M25 can revert by super-suppressor mutations in unlinked genes, thus confirming previous evidence that M54 contains a nonsense codon. At least three new super suppressors of M54 have been detected. All four super suppressors (including one previously detected) when combined with M54 result in high levels of all five of the arom enzymic activities in the form of arom multienzyme complexes very similar to (but not necessarily identical with) that in wild type (WT).-Evidence has also been obtained that the two non-complementing mutants can yield revertants which appear to result from true back mutations and produce arom aggregates essentially indistinguishable from that of WT. In addition, M25, but not M54, when plated on quinic acid yields revertants (secondary mutants) some of which are phenotypically indistinguishable from arom-2 primary mutants and others of which, although also mapping within the arom-2 gene, exhibit unusual properties. Genetic evidence indicates that the M25 secondary mutants are localized within the arom-2 gene, but that they arise from mutational events more complex than ones resulting in single base pair changes in the M25 codon.-The recovery of secondary arom-2 mutants as revertants of non-complementing arom mutants provides strong evidence, independent of earlier recombination data, that non-complementing arom mutants are located within the arom-2 structural gene of the arom gene cluster. In addition, the occurrence and characteristics of these secondary arom-2 mutants provide strong evidence, independent of the results with nonsense suppressors, that the arom gene cluster is transcribed, beginning with the arom-2 gene, as a single polycistronic messenger ribonucleic acid (mRNA) molecule which is subsequently translated into the arom multienzyme complex.     

Mutations that confer resistance to 2-deoxyglucose reduce the specific activity of hexokinase from Myxococcus xanthus

The glucose analog 2-deoxyglucose (2dGlc) inhibits the growth and multicellular development of Myxococcus xanthus. Mutants of M. xanthus resistant to 2dGlc, designated hex mutants, arise at a low spontaneous frequency. Expression of the Escherichia coli glk (glucokinase) gene in M. xanthus hex mutants restores 2dGlc sensitivity, suggesting that these mutants arise upon the loss of a soluble hexokinase function that phosphorylates 2dGlc to form the toxic intermediate, 2-deoxyglucose-6-phosphate. Enzyme assays of M. xanthus extracts reveal a soluble hexokinase (ATP:D-hexose-6-phosphotransferase; EC 2.7.1.1) activity but no phosphotransferase system activities. The hex mutants have lower levels of hexokinase activities than the wild type, and the levels of hexokinase activity exhibited by the hex mutants are inversely correlated with the ability of 2dGlc to inhibit their growth and sporulation. Both 2dGlc and N-acetylglucosamine act as inhibitors of glucose turnover by the M. xanthus hexokinase in vitro, consistent with the finding that glucose and N-acetylglucosamine can antagonize the toxic effects of 2dGlc in vivo.     

Isolation and characterization of Saccharomyces cerevisiae mutants deficient in S-adenosylmethionine decarboxylase, spermidine, and spermine.

Four mutants were isolated from Saccharomyces cerevisiae that are deficient in S-adenosylmethionine decarboxylase (spe2). All four mutants are chromosomal and fall into a single complementation group tightly linked to arg1. Since one of the mutants contained a temperature-sensitive activity, this complementation group defines the structural gene. Mutants totally lacking enzymic activity did not contain spermidine or spermine and had a greatly increased doubling time when grown in the absence of these two polyamines. Addition of 10(-6) M spermidine or 10(-5) M spermine, but not putrescine or cadaverine, restored the doubling time to that of the wild type. Diploids formed from a cross of two mutants completely deficient in spermidine and spermine were unable to sporulate in the absence of added spermidine or spermine. We obtained evidence that arg1 was not located on any of the 17 known chromosomes, and therefore we postulate that arg1 and spe2 are located on a new 18th chromosome.