Genetic Analysis and Molecular Mapping of an All-Stage Stripe Rust Resistance Gene in Triticum aestivum-Haynaldia villosa Translocation Line V3

Triticum aestivum-Hayaldia villosa translocation line V3 has shown effective all-stage resistance to the seven dominant pathotypes of Puccinia striiforms f.sp.tritici prevalent in China.To elucidate the genetic basis of the resistance,the segregating populations were developed from the cross between V3 and susceptible genotype Mingxian 169,seedlings of the parents and F 2 progeny were tested with six prevalent pathotypes,including CYR29,CYR31,CYR32-6,CYR33,Sun11-4,and Sun11-11,F 1 plants and F 3 lines were also inoculated with Sun11-11 to confirm the result further.The genetic studied results showed that the resistance of V3 against CYR29 was conferred by two dominant genes,independently,one dominant gene and one recessive gene conferring independently or a single dominant gene to confer resistance to CYR31,two complementary dominant genes conferring resistance to both CYR32-6 and Sun11-4,two independently dominant genes or three dominant genes（two of the genes show cumulative effect） conferring resistance to CYR33,a single dominant gene for resistance to Sun11-11.Resistance gene analog polymorphism（RGAP） and simple-sequence repeat（SSR） techniques were used to identify molecular markers linked to the single dominant gene（temporarily designated as YrV3） for resistance to Sun11-11.A linkage map of 2 RGAP and 7 SSR markers was constructed for the dominant gene using data from 221 F 2 plants and their derived F 2：3 lines tested with Sun11-11 in the greenhouse.Amplification of the complete set of nulli-tetrasomic lines of Chinese Spring with a RGAP marker RG1 mapped the gene on the chromosome 1B,and then the linked 7 SSR markers located this gene on the long arm of chromosome 1B.The linkage map spanned a genetic distance of 25.0 cM,the SSR markers Xgwm124 and Xcfa2147 closely linked to YrV3 with genetic distances of 3.0 and 3.8 cM,respectively.Based on the linkage map,it concluded that the resistance gene YrV3 was located on chromosome arm 1BL.Given chromosomal location,the reaction patterns and pedigree analysis,YrV3 should be a novel gene for resistance to stripe rust in wheat.These closely linked markers should be useful in stacking genes from different sources for wheat breeding and diversification of resistance genes against stripe rust.     

Combining Phytate/Ca2+ Fractionation with Trichloroacetic Acid/Acetone Precipitation Improved Separation of Low-Abundant Proteins of Wheat (Triticum aestivum L.) Leaf for Proteomic Analysis

Proteomic assessment of low-abundance leaf proteins is hindered by the large quantity of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) present within plant leaf tissues. In the present study, total proteins were extracted from wheat (Triticum aestivum L.) leaves by a conventional trichloroacetic acid (TCA)/acetone method and a protocol first developed in this work. Phytate/Ca²⁺ fractionation and TCA/acetone precipitation were combined to design an improved TCA/acetone method. The extracted proteins were analysed by two-dimensional gel electrophoresis (2-DE). The resulting 2-DE images were compared to reveal major differences. The results showed that large quantities of Rubisco were deleted from wheat leaf proteins prepared by the improved method. As many as (758±4) protein spots were detected from 2-DE images of protein extracts obtained by the improved method, 130 more than those detected by the TCA/acetone method. Further analysis indicated that more protein spots could be detected at regions of pI 4.00–4.99 and 6.50–7.00 in the improved method-based 2-DE images. Our findings indicated that the improved method is an efficient protein preparation protocol for separating low-abundance proteins in wheat leaf tissues by 2-DE analysis. The proposed protocol is simple, fast, inexpensive and also applicable to protein preparations of other plants.     

基因枪介导的转PYL5基因小麦的获得与鉴定

【目的】通过基因枪介导共转化,获得转拟南芥抗旱基因PYL5的小麦植株,为转基因小麦的抗旱功能研究奠定基础。【方法】以小麦品种西农889、绵阳19和宁春16为供试材料,通过基因枪介导法将诱导型启动子Rd29A启动的PYL5基因和筛选标记基因Bar共转化到小麦幼胚愈伤组织中,经过除草剂PPT(Phosphinothricin)筛选和愈伤组织分化,获得再生植株。根据目标基因PYL5和Bar基因序列分别设计特异引物,对移栽成活的小麦T0代再生植株进行基因特异性PCR检测。【结果】采用基因枪分别轰击西农889的1800个、绵阳19的800个和宁春16的800个幼胚愈伤组织,经过筛选和分化分别获得了9,5和14株小麦再生植株;对转基因小麦T0代再生植株的基因特异性PCR检测结果表明,西农889、绵阳19和宁春16 Bar基因的转化率分别为0.280%,0.500%和0.750%,PYL5基因的转化率分别为0.110%,0.125%和0.500%,Bar和PYL5基因的共转化率分别为0.110%,0.125%和0.500%。【结论】PYL5基因成功转入到了小麦品种西农889、绵阳19和宁春16中。     

Establishment of an efficient regeneration and genetic transformation system for Malus prunifolia Borkh. ‘Fupingqiuzi’

Malus prunifolia Borkh. ‘Fupingqiuzi’ has significant ecological and economic value and plays a key role in germplasm development and resistance research.  However, its long juvenile phase and high heterozygosity are barriers to the identification of ‘Fupingqiuzi’ progeny with excellent traits.  In-vitro regeneration techniques and Agrobacterium-mediated genetic transformation systems can efficiently produce complete plants and thus enable studies of gene function.  However, optimal regeneration and genetic transformation systems for ‘Fupingqiuzi’ have not yet been developed.  Here, we evaluated the factors that affect the in-vitro regeneration and transformation of ‘Fupingqiuzi’.  The best results were obtained when transverse leaf sections were used as explants, and they were grown in dark culture for three weeks with their adaxial sides contacting the culture medium (MS basal salts, 30 g L⁻¹ sucrose, 8 g L⁻¹ agar, 5 mg L⁻¹  6-benzylaminopurine (6-BA), 2 mg L⁻¹ thidiazuron (TDZ), and 1 mg L⁻¹ 1-naphthlcetic acid (NAA), pH 5.8).  A genetic transformation system based on this regeneration system was optimized: after inoculation with A. tumefaciens solution for 8 min, 4 days of co-culture, and 3 days of delayed culture, the cultures were screened with cefotaxime (150 mg L⁻¹) and kanamycin (15 mg L⁻¹).  We thus established an efficient regeneration and genetic transformation system for ‘Fupingqiuzi’, enabling the rapid production of transgenic material.  These findings make a significant contribution to apple biology research     

4个小麦—柔软滨麦草易位系苗期抗条锈基因的遗传分析

旨在开发和利用柔软滨麦草的基因,丰富小麦抗条锈基因库。利用小麦条锈菌流行小种CYR32和CYR33对M851-1、M8724-1、M8725-2和M8657-2 4个小麦-柔软滨麦草易位系进行苗期抗条锈性遗传分析。结果表明,M851-1对CYR32的抗条锈性由1对隐性基因控制;M8724-1对CYR32的抗条锈性由2对隐性基因独立作用控制,对CYR33的抗条锈性由1对隐性基因控制;M8725-2对CYR32的抗条锈性由2对显性基因互补作用控制,对CYR33的抗条锈性由1显1隐2对基因独立控制;M8657-2对CYR32的抗条锈性由1对隐性基因控制,对CYR33的抗条锈性由2对显性基因独立作用控制。研究结果初步明确这4个小麦-柔软滨麦草易位系抗条锈性遗传规律,有助于进一步利用这些易位系进行小麦抗条锈病育种。     

糜子感染黑穗病菌后的生理变化及与抗病性关系

为明确糜子对黑穗病的抗性生理机制,在大田条件下,以不同抗性水平的10个糜子品种为材料,采用种子饱和接种法人工接种黑粉菌,测定了三叶期不同糜子品种叶片超氧化物歧化酶(SOD)、过氧化物酶(POD)、β-1,3-葡聚糖酶、几丁质酶活性以及丙二醛(MDA)含量。结果表明,人工接种黑粉菌后不同抗性水平糜子品种叶片SOD、β-1,3-葡聚糖酶和几丁质酶活性有所提高,最大增幅分别达92%、16%和31%;MDA含量有所增加,最大增幅达20%;POD活性呈不同变化。人工接种条件下抗性水平较高品种的保护酶系统酶活性以及病程相关蛋白活性提升幅度较大,MDA含量增加幅度较小。各生理指标与抗病性的相关分析表明,SOD、POD、β-1,3-葡聚糖酶以及几丁质酶活性与发病率呈极显著负相关,相关系数分别为0.941、0.468、0.514和0.757;MDA含量与发病率呈显著正相关,相关系数为0.378。     

Occurrence of ‘Candidatus Phytoplasma ziziphi’ in apple trees in China

Young apple trees exhibiting symptoms of little leaf, margin involute and yellows were observed in an open‐air nursery in Yangling, Shaanxi, China. Transmission electron microscopy showed typical phytoplasma bodies in the sieve tube elements of symptomatic leaf samples. In two nested polymerase chain reaction (PCR) assays, products of expected size of 1.1 and 1.2 kb were separately generated from the total DNAs of symptomatic samples. A restriction fragment length polymorphism (RFLP) analysis of the purified 1.2 kb PCR products indicated that the disease associated with apple trees was ‘Ca. Phytoplasma ziziphi’. To our knowledge, this is the first report of ‘Ca. Phytoplasma ziziphi’ infecting apple trees in China.     

不同杀菌剂对烟草白粉病菌的室内毒力及田间防效

【目的】比较不同杀菌剂对烟草白粉病的室内毒力和田间药效,以筛选出适合于治疗烟草白粉病的杀菌剂。【方法】分别采用孢子萌发法和田间防效试验,研究30%醚菌酯悬浮剂、15%三唑酮可湿性粉剂、50%百菌清可湿性粉剂、50%退菌特可湿性粉剂、25%乙嘧酚悬浮剂和对照药剂70%甲基硫菌灵可湿性粉剂对烟草白粉病的室内毒力及田间防治效果。【结果】室内毒力测定结果和田间药效试验结果表明,30%醚菌酯悬浮剂对烟草白粉病菌孢子萌发的EC50为2.29mg/L,其3 000倍液的防治效果可达84.29%;25%乙嘧酚悬浮剂对烟草白粉病菌孢子萌发的EC50为2.88mg/L,其1 000倍液的防治效果可达81.57%;对照药剂75%甲基硫菌灵可湿性粉剂对烟草白粉病菌孢子萌发的EC50为15.49mg/L,其1 000倍液的防治效果只有31.76%;其他杀菌剂的防治效果均低于醚菌酯和乙嘧酚,但高于对照药剂甲基硫菌灵。【结论】醚菌酯和乙嘧酚均能有效地防治烟草白粉病,且效果优于对照药剂甲基硫菌灵,为生产提供了更高效可供替代的化学药剂品种。     

小偃6号成株期高温抗条锈性遗传分析

为揭示小偃6号抗病机制和培育持久抗病品种,采用常规杂交分析方法,在小麦抽穗期利用小麦条锈菌小种CYR30、CYR32和Su11-4对小偃6号、铭贤169及其杂交F1、F2、F2∶3接种,平均气温达到21℃时对小偃6号进行了抗条锈性调查和遗传分析。结果显示,接种CYR30、CYR32时,F1代表现高感,F2代群体中抗感分离比例符合1 R∶15 S的理论比例。接种Su11-4时,F1代表现高抗,F2代群体中抗感分离比例符合3R∶1S的理论比例。研究表明小偃6号对CYR30、CYR32的抗病性均由2对隐性基因累加作用控制,对Su11-4的抗病性由1对显性基因控制。     

碳同位素分辨率与小麦光合生理指标的关系

为了解碳同位素分辨率(Δ)与小麦光合生理指标的关系,以洛旱6号和西农389的154个F4代株系中的24个高Δ株系和24个低Δ株系及亲本为供试材料,分析了陕西杨凌和永寿两种不同雨养环境条件下小麦籽粒Δ与灌浆期叶片光合生理指标的相关性,并对高、低Δ材料间光合生理指标和产量进行了比较分析。结果表明,在降雨较多的杨凌地区,高Δ株系表现出高的籽粒产量;在降雨较少、相对干旱的永寿地区,高、低Δ株系间籽粒产量差异不显著,但低Δ株系表现出低的气孔导度。在杨凌生态条件下,灌浆中期籽粒Δ与叶片蒸腾速率、光合速率、气孔导度和Ci/Ca生理指标的相关性在小麦整个灌浆期最为显著;在永寿生态条件下,灌浆中期籽粒Δ与蒸腾速率和Ci/Ca均相关显著。说环境条件影响小麦籽粒Δ与光合生理指标的关系。