Disruption of Energy Metabolism and Reactive Oxygen Species Homeostasis in Honglian Type-Cytoplasmic Male Sterility(HL-CMS)Rice Pollen

Honglian type-cytoplasmic male sterility(HL-CMS)is caused by the inter-communication between the nucleus and mitochondria.However,the mechanisms by which sterility genes regulate metabolic alterations and changes in mitochondrial morphology in the pollen of HL-CMS remain unclear.In this study,we compared the morphological differences between the pollen of the male sterile line YA and the near-isogenic line NIL-Rf6 using hematoxylin-eosin staining and 4ʹ,6-diamidino-2-phenylindole(DAPI)staining.HL-CMS is characterized by gametophytic sterility,where the aborted pollen grains are empty,and the tapetal layer remains intact.Transmission electron microscopy was employed to observe mitochondrial morphological changes at the microspore stage,revealing significant mitochondrial alterations,characterized by the formation of'large spherical mitochondria',occurred at the binucleate stage in the YA line.Additionally,metabolomics analysis revealed decreased levels of metabolites associated with the carbohydrate and flavonoid pathways.Notably,the decrease in flavonoids was found to contribute to an elevation in reactive oxygen species(ROS)levels.Therefore,we propose a model in which rice fertility is modulated by the levels of pollen carbohydrates and flavonoid metabolites,with impaired mitochondrial energy production and reduced flavonoid biosynthesis as the main causes of ROS accumulation and pollen abortion in rice.

Dynamically changing antineutrophil cytoplasmic antibodies in granulomatosis with polyangiitis:A case report

BACKGROUND Granulomatosis with polyangiitis(GPA)is one of the most prevalent forms of the antineutrophil cytoplasmic antibody(ANCA)-associated vasculitis.GPA is characterized histologically by necrotizing granulomatous inflammation in addition to vasculitis.The diagnosis of GPA depends on clinical presentation,serological evidence of a positive ANCA,and/or histological evidence of necrotizing vasculitis or granulomatous destructive parenchymal inflammation.Cytoplasmic ANCA(c-ANCA)is positive in 65%-75% of GPA patients,accompanied by proteinase 3(PR3),the main target antigen of c-ANCA,another 5% of GPA patients had negative ANCA.CASE SUMMARY The patient,a 52-year-old male,presented with unexplained nasal congestion,tinnitus,and hearing loss.After a duration of 4 months experiencing these symptoms,the patient subsequently developed fever and headache.The imaging examination revealed the presence of bilateral auricular mastoiditis and partial paranasal sinusitis,and the ANCA results were negative.The anti-infective therapy proved to be ineffective,but the patient's symptoms and fever were quickly relieved after 1 wk of treatment with methylprednisolone 40 mg once a day.However,after continuous use of methylprednisolone tablets for 3 months,the patient experienced a recurrence of fever accompanied by right-sided migraine,positive c-ANCA and PR3,and increased total protein in cerebrospinal fluid.The and cyclophosphamide 0.8 g monthly,the patient experienced alleviation of fever and headache.Additionally,the ANCA levels became negative and there has been no recurrence.CONCLUSION For GPA patients with negative ANCA,there is a potential for early missed diagnosis.The integration of histopathological results and multidisciplinary communication plays a crucial role in facilitating ANCA-negative GPA.

Synergy in Rice Immunity:Exploring Strategies of Coordinated Disease Defense Through Receptor-Like Kinases and Receptor-Like Cytoplasmic Kinases

Receptor-like kinases(RLKs)and receptor-like cytoplasmic kinases(RLCKs)play an indispensable role in the perception and transmission of extracellular signals in plants.In rice,these kinases actively participate in immune responses against a variety of pathogens,including fungi,bacteria,and viruses.However,research on the specific response mechanisms and the spectrum of different kinase activities against various pathogens remains insufficient.This review provides an in-depth and comprehensive overview of the types and functions of RLKs and RLCKs involved in disease resistance,emphasizing the central role of certain RLKs and RLCKs in the plant immune system.These kinases can recognize specific molecular patterns of pathogens and rapidly initiate an immune response in rice.Furthermore,the activity and functional regulation of these key kinases are tightly controlled by various post-translational modifications,such as phosphorylation and ubiquitination.This meticulous regulation ensures that the rice immune system's response is both precise and timely,effectively balancing the intensity of the immune response and preventing potential issues caused by either hyperactivity or insufficiency.By synthesizing current research findings,this review not only broadens our understanding of the role of RLKs and RLCKs in plant immunity but also provides new perspectives and strategies for future research on disease resistance breeding in rice.Future studies are expected to delve deeper into the signaling networks and regulatory mechanisms of these kinases,exploring their potential in agricultural production to develop rice varieties with enhanced disease resistance.

Morphological and cytological assessments reveal pollen degradation causes pollen abortion in cotton cytoplasmic male sterility lines

Background Understanding the mechanism of male sterility is crucial for producing hybrid seeds and developing sterile germplasm resources.However,only a few cytoplasmic male sterility(CMS)lines of cotton have been produced due to several challenges,like inadequate variation of agronomic traits,incomplete sterility,weak resilience of restorer lines,and difficulty in combining strong dominance.Therefore,the morphological and cytological identification of CMS in cotton will facilitate hybrid breeding.Results Two F_(2) segregating populations of cotton were constructed from cytoplasmic male sterile lines(HaA and 01A,maternal)and restorer lines(HaR and 26R,paternal).Genetic analysis of these populations revealed a segregation ratio of 3:1 for fertile to sterile plants.Phenotypic analysis indicated no significant differences in traits of flower bud development between sterile and fertile plants.However,sterile plants exhibited smaller floral organs,shortened filament lengths,and anther atrophy on the flowering day in comparison with the fertile plants.When performed scanning electron microscopy(SEM),the two F_(2) populations revealed morphological variations in the anther epidermis.Cellular analysis showed no significant differences in pollen development before pollen maturation.Interestingly,between the pollen maturation and flowering stages,the tapetum layer of sterile plants degenerated prematurely,resulting in abnormal pollen grains and gradual pollen degradation.Conclusion The results of this study suggest that fertility-restoring genes are controlled by a single dominant gene.Sterile plants exhibit distinctive floral morphology,which is characterized by stamen atrophy and abnormal anthers.Pollen abortion occurs between pollen maturity and flowering,indicating that premature tapetum degradation may be the primary cause of pollen abortion.Overall,our study provides a theoretical basis for utilizing CMS in hybrid breeding and in-depth investigation of the dominant configuration of cotton hybrid combinations,mechanisms of sterility,and the role of sterile and restorer genes.

Potential regulatory mechanism and clinical significance of synaptotagmin binding cytoplasmic RNA interacting protein in colorectal cancer

BACKGROUND Colorectal cancer(CRC)causes many deaths worldwide.Synaptotagmin binding cytoplasmic RNA interacting protein(SYNCRIP)is an RNA-binding protein that plays an important role in multiple cancers by epigenetically targeting some genes.Our study will examine the expression,potential effect,biological function and clinical value of SYNCRIP in CRC.AIM To examine the expression,potential effect,biological function and clinical value METHODS The expression of SYNCRIP was examined by immunohistochemistry arrays and high-throughput data.The effect of SYNCRIP gene in CRC cell growth was evaluated by CRISPR-Cas9 technology.The target genes of SYNCRIP were calculated using various algorithms,and the molecular mechanism of SYNCRIP in CRC was explored by mutation analysis and pathway analysis.The clinical value of SYNCRIP in prognosis and radiotherapy was revealed via evidence-based medicine methods.RESULTS The protein and mRNA levels of SYNCRIP were both highly expressed in CRC samples compared to nontumorous tissue based on 330 immunohistochemistry arrays and 3640 CRC samples.Cells grew more slowly in eleven CRC cell lines after knocking out the SYNCRIP gene.SYNCRIP could epigenetically target genes to promote the occurrence and development of CRC by boosting the cell cycle and affecting the tumor microenvironment.In addition,CRC patients with high SYNCRIP expression are more sensitive to radiotherapy.CONCLUSION SYNCRIP is upregulated in CRC,and highly expressed SYNCRIP can accelerate CRC cell division by exerting its epigenetic regulatory effects.In addition,SYNCRIP is expected to become a potential biomarker to predict the effect of radiotherapy.

Murtagh安全诊断策略联合思维导图构建临床思维在全科教学门诊中的应用

全科教学门诊是培养住院医师规范化培训(简称住培)全科医生临床思维、提升全科诊疗能力的重要培训方法。本文用1例以乏力为主要表现的患者为教学案例,介绍全科教学门诊中采用澳大利亚著名全科医学专家John Murtagh提出的Murtagh安全诊断策略启发住培全科医生对乏力进行诊断与鉴别诊断:(1)引起乏力的常见疾病有哪些?(2)哪些重要疾病是不能忽视的?(3)乏力有什么容易被遗漏的疾病?(4)是否存在潜在的容易被掩盖的疾病?(5)患者是否有话没有说?结合病史、体格检查和实验室检查结果初步诊断为抗中性粒细胞胞浆抗体相关性血管炎引起急进性肾小球肾炎可能性大,及时转诊,肾病理检查确诊乏力的病因为抗中性粒细胞胞浆抗体相关性肾小球肾炎,取得满意疗效。带教老师基于Murtagh安全诊断策略帮助住培全科医生构建乏力鉴别、分析的系统知识框架,从而提升住培全科医生的临床逻辑思维能力和解决临床实际问题的能力;以思维导图为辅助工具,将Murtagh安全诊断策略诊断与鉴别诊断思路可视化、形象化,优化教学效果。

Influential Factors of Restorer of New Cytoplasmic Male Sterile(NER) on Anther Culture

Some influential factors of anther culture were studied preliminarily by conducting anther culture of the restorers of new cytoplasmic male sterile （NER）. Several results were obtain from this experiment and they were listed as follow：① MS cultrure medium with such hormones as 2,4-D 2 mg/L,6-BA 0.5 mg/L, NAA 0.5 mg/L was the best suitable for callus induction of NER. ②The difference of induction rate was significantly different between different plant age groups. From the 110th day to 141th day,the induction rate was increased with the increase of age and the difference of induction rate reached 0.01 significant difference level. The induction rate reached the highest value in the 141th day then it declined gradually. ③The combined use of 2, 4-D and 6-BA with proper increase of 2,4-D was good for inducing callus. ④The green plantlet induction rate of NER was increased when the concentration of 6-BA increased from 2 mg/L to 4 mg/L. Adding ZT from 0.5 mg/L to 2 mg/L. 6-BA would led 2.47% increase of green plantlet olantlet induction rate.

RAPD and ISSR Markers of Fertility Restoring Gene for Aegilops kotschyi Cytoplasmic Male Sterility in Wheat

LK783 was found to be a good fertility restorer for K-type male sterility of wheat (Triticum aestivum L.). RAPD and ISSR (inter-simple sequence repeat polymorphism) markers were employed to map the major restoring gene in LK783. Maintainer and restorer DNA pools were established using the extreme sterile and fertile plants among KJ5418A//911289/LK783 F 1 population, respectively. Four hundred and eighteen RAPD primers and 33 ISSR primers were used for screening polymorphisms between the two pools, and amplification bands using a RAPD primer of OPK18 and an ISSR primer of UBC-845 were found polymorphic between the two pools. Linkage analysis showed that OPK18 450 and UBC-845 800 were linked to the restoring gene in LK783. The distance between the restoring gene and OPK18 450 was (15.07±6.28) cM (centiMorgan), with the distance between the restoring gene and UBC-845 800 being (8.20±4.85) cM. The marker of UBC-845 800 was located on chromosome 1BS by amplifying nulli-tetrasomics and 1B ditelosomics of Chinese Spring with the primer of UBC-845, indicating that the restoring gene in LK783 was located on 1BS. The breeding for new fertility restorer lines of K-type cytoplasmic male sterility of wheat would be facilitated by using the two markers.

Studies on Contamination of Cytoplasm DNA and Its Control in Plant Chromosone Microdissection

染色体微切割和微克隆已成为复杂基因组研究的有效途径 ,但是操作过程中的核外DNA的污染一直是令人担心的问题。通过研究植物染色体微切割 (微分离 )和微切割的染色体DNA扩增过程中细胞质DNA的污染问题 ,表明目前常用的植物染色体微切割过程中 ,细胞质DNA的污染几乎难以避免 ,并提出了一个改进的降低细胞质DNA污染的方法 ,对如何控制细胞质DNA的污染进行了详细的讨论。

AFLP Analysis of Cytoplasmic Male-sterile Line and Its Maintainer Line of Pepper

[Objective] The aim of this study was to analyze the cytoplasmic male sterile line 21A and its maintainer line 21B of peppers by AFLP,and lay the foundation for further studies on molecular mechanism of the cytoplasmic male sterility in peppers.[Method] Cytoplasmic male sterility(CMS)line 21A and its maintainer line 21B were analyzed by AFLP to obtain the specific amplified fragments of cytoplasmic male sterile line 21A,while the specific amplified fragments were recovered or sequenced,and analyzed by BLAST...

Microtubule Structure and Male Sterility in a Gene-Cytoplasmic Male Sterile Line of Rice, Zhen Shan 97A

Histological changes that occur during microsporogenesis are documented in a gene-cytoplasmic male sterile rice ( Oryza saliva L.) line, Zhen Shan 97A, its maintainer line, Zhen Shan 97B, and the restorer line, Ce64 of a Mine hybrid rice production system. In the restorer line, Ce64, the developing microsporocytes have dense cytoplasm and a distinct set of circumferential microtubules around the nucleus. Successive cytokinesis results in the formation of tetrads. The microtubules within the cells of tetrads and microspores radiate from the surface of the nucleus towards the outer edge of the cytoplasm. Subsequent pollen development is normal. During the course of microspore formation tubulin speckles can be found in the cytoplasm. The general pattern of development and microtubule organization in the maintainer lined Zhen Shan 97B, is similar to Ce64, except that a few more tubulin speckles appear during microspore formation. In the case of the mate sterile line, Zhen Shan 97A, a number of abnormalities can be discerned during early microsporogenesis. These include vacuoles forming within the developing microsporocyte and faintly stained microtubules with no defined distribution pattern. Prominent tubulin speckles are common within the cytoplasm. For those microsporocytes that undergo meiosis, no defined organizational patterns of microtubules can be found within the tetrad. All microspores abort soon after. Abnormalities and defects in microtubule organization observed in Zhen Shan 97A showed that complex interactions between the cytoplasm and the nucleus began at very early stage of microsporocyte development.

Preliminary Studies on the Effect of Photoperiod and Temperature on Male Fertility of a Cytoplasmic Male Sterile Line in Wheat

[Objective] The study aimed to reveal the effect of photoperiod and temperature on male fertility of cytoplasmic male sterile line Vtai911289a in wheat and discuss the mechanism of male fertility alteration. [ Method ] The sowing-date tests and designed conditions were conducted during 2003 -2005. [ Result] Fertility of Vtai911289a, could alter under specific photoperiod and temperature conditions. Temperature is one of the main factors influencing male fertility of the male sterile lines. Vtai911289a, showed stable sterility under the condition of the mean of daily temperature at fertility sensitive stage lower than 19℃ and presented partial fertility when the mean of daily temperature at fertility sensitive stage lower than 20 - 22℃. Photoperiod to some extent affects the male fertility of Vtai911289a, long-day condition is helpful for the male fertility of the sterile line. [ Conclusion] The application of photoperiod temperature-sensitive cytoplasmic male sterile line in production has a higher safety than that of temperature sensitive sterile line.

辣椒胞质雄性不育恢复基因CAPS标记的PARMS转换及其应用

Co1Mod1-CAPS是一个可广泛应用于筛选辣椒胞质雄性不育恢复源的CAPS分子标记。为了将其转化为可高通量检测的分子标记以提高选择效率,本研究以Co1Mod1-CAPS标记扩增10份不同类型的辣椒三系材料,分析其扩增产物在不育系、保持系和恢复系材料间的序列差异,从而基于序列差异将其转化为PARMS高通量检测标记并对321份辣椒种质材料进行恢复源筛选。结果表明,在Co1Mod1-CAPS标记扩增产物第601个碱基处存在一个T→C碱基的变异,即不育系和保持系材料在该处的碱基均为T,而恢复系材料在该处的碱基为C,后基于该碱基变异成功将Co1Mod1-CAPS转化为高通量标记Co1Mod1-PARMS,并在321份种质材料中筛选出146份资源携带纯合恢复基因RfRf,其中小果形且辣味重的辣椒类型携带RfRf基因型的个体占比高,指形椒的占比最高,为59.5%,樱桃椒占比次之,为56.5%,甜椒类型辣椒(灯笼椒)携带RfRf基因型的个体占比最低,为8.3%。该研究结果与前人研究较为一致,表明该标记有效。因此,Co1Mod1-PARMS高通量检测标记可应用于辣椒胞质雄性不育恢复系的分子标记辅助选育,从而促使胞质雄性不育恢复系选育效率的提升。

大肠杆菌胞质非特异性二肽酶对γ-氨基丁酸的催化作用研究

γ-氨基丁酸(γ-aminobutyric acid,GABA)广泛应用于食品和医药等领域,目前主要利用大肠杆菌(Escherichia coli)作为宿主菌,由酶催化方法生产。在全细胞转化过程中,GABA会进入三羧酸循环进行降解,导致转化率低下;通过敲基因阻碍此途径又会严重影响宿主菌的生长。该研究揭示了一条新的GABA的消耗途径:GABA在胞质非特异性二肽酶(cytosolic nonspecific dipeptidase,pepD)的作用下缩合形成二肽。该文构建了2个工程菌株:敲除E.coli BL21(DE3)基因组上的pepD基因,构建了ΔpepD菌株;在E.coli BL21(DE3)中过表达了pepD基因,构建了OE pepD菌株。以GABA和谷氨酸钠(monosodium glutamate,MSG)为底物,进行全细胞转化反应。结果表明,该酶不能催化MSG发生反应,但是能催化GABA发生反应。通过研究2个菌株消耗GABA的历程,发现OE pepD的GABA消耗量比ΔpepD菌株高出17.5%,消耗量在2 h之后都不再增加。利用LC-MS鉴定纯酶催化反应的产物,证实产物是GABA形成的二肽。该研究构建的菌株可以在不影响大肠杆菌生长的情况下有效提高GABA的产量,为GABA的工业化生产提供有益参考。

Cytological Study on Zidao Type Cytoplasmic Male Sterile Line-Yingxiang A

Anthers of male sterile line and its maintainer line of Zidao type rice, Yingxiang A and Yingxiang B were studied by cytological methods, such as F-KI staining, silver nitrate staining and paraffin sections, It is found that in Yingxiang A, there are some abnormalities in meiosis, represented by unequal division at the meiotic phase; the abortion of CMS line arises at the meiotic phase and is almost completed at the phase of monomicrospore, but only a tiny amount of aborted mi- crospores are observed; the abortion is complete and thorough, and none normal functional pollens are found; the morphology of aborted pollens is similar to that of typically aborted pollens; the tapetal cells still exist in anthers from the microsporocyte stage to the tetrad stage, but they are abnormally hyperplastic. However, in Yingxiang B, there are no abnormalities in meiosis at the meiotic phase; the tapetal cells exist in anthers before the tetrad stage, but they disappear completely when the pollen are mature. It is indicated that the abortion of CMS Yingxiang line mainly occurs at the meiotic phase, and is basically completed at the phase of monomicrospore. The abortion is probably caused by the abnormally hyperplastic tapetal cells. The abnormalities in tapetal cells may affect the normal development of pollens, resulting in the emergence of abortion. In addition, the abortion in Yingxiang A is complete and thorough, which is of great significance in hybrid breeding.

甲巯咪唑诱发中性粒细胞胞质抗体阳性相关性血管炎的文献分析

目的探讨甲巯咪唑诱发中性粒细胞胞质抗体阳性(ANCA)相关性血管炎发生情况和临床特点,为临床合理用药提供参考。方法检索万方数据库、中国知网、PubMed、Web of Science数据库,收集甲巯咪唑致ANCA相关性血管炎的个案报道,检索时限为从建库起至2023年10月31日,描述性分析患者人口学特征、药物使用情况、并发症、治疗及转归。结果共纳入文献14篇,合计患者14例,其中男3例,女11例,年龄8~79岁,平均年龄为(47.79±23.47)岁。4例在服药1年内发生,9例在服药2~12年内发生,1例在服药后24年发生。ANCA相关性血管炎累及肾脏8例、累及皮肤和肺部5例、累及视力及累及心脏各2例。所有患者均停用甲巯咪唑,2例患者未经治疗自动好转,1例患者经过抗感染后好转,其他病例均接受糖皮质激素或免疫抑制剂治疗。1例患者死亡,其他患者经过治疗后均好转或治愈。结论甲巯咪唑诱发的ANCA相关性血管炎较为罕见,患者起病时间较长,主要累及皮肤、肾脏、肺部、视力等多系统/器官,临床医生应注意药源性ANCA相关性血管炎与原发性血管炎相鉴别,及时停药。出现严重脏器损伤时,及时予以糖皮质激素及免疫抑制剂治疗,避免病情加重危及生命。

Preliminary Study on Transgenesis by Injecting Exogenous DNA into Zygote Cytoplasm of Buffalo

[Objective] This study aimed to investigate the feasibility of transgenesis by injecting exogenous DNA into zygote cytoplasm of Buffalo. [Method] Buffalo oocytes were randomly divided into two groups 20-22 h after in vitro maturation. One group of oocytes was introduced with about 7.5 pl of 50 μg/ml DNA solution containing linear EGFP fragment by cytoplasmic injection 7-10 h or 18-20 h after in vitro fertilization （IVF）; the other group of oocytes was introduced with mixture of a single buffalo sperm and about 7.5 pl of 50 μg/ml DNA solution containing linear EGFP fragment by cytoplasmic injection （generally called ICSI-Mediated Gene Transfer, ICSI-Tr）. Expression of exogenous DNA was observed and recorded during the process of embryonic development. [Result] Early embryonic gene expression efficiency and blastocyst gene expression efficiency in IVF injection group showed no significant difference compared with that in ICSI-Tr group （P0.05）. In addition, the cleavage rate and early embryonic gene expression efficiency in IVF injection group were significantly higher with injection at 7-10 h post IVF than that at 18-20 h post IVF （P0.05）. [Conclusion] These results indicate that transgenic buffalo embryos can be generated by injecting exogenous DNA into cytoplasm of IVF oocytes, and the optimal injection time is 7-10 h post IVF.

Improvement of in vitro Cytoplasmic Maturation of Denuded Porcine Oocytes by Coculture with Follicular Mural Granulosa Cells

[Objective] This study aimed to improve the in vitro maturation quality of denuded porcine oocytes and provide scientific basis for establishing a stable and efficient denuded oocyte culture system. [Method] The first polar body extrusion rate, oocyte glutathione （GSH） content, positive rate of brilliant cresyl blue （BCB） staining and development potential of activated oocytes or fertilized oocytes were employed as main indicators to investigate the effects of follicular mural granulosa cell （MGC） coculture on cytoplasmic maturation of cumulus cell-removal oocytes （Denuded Oocyte, DO）. [Result] According to in vitro maturation results, compared with DO group, the first polar body extrusion rate of porcine oocytes in DO＋MGC group was not significantly different, but the nuclear maturation process was improved and was more similar to that in COC （cumulus-oocyte complex） group. Detection of GSH content in mature oocytes showed that there was no significant difference between DO＋ MGC group （optical density of 1 053.67） and COC group （optical density of 1 426.00） or between DO＋MGC group and COC＋GC group （optical density of 1 541.00）, however, GSH content in mature oocytes of DO group （optical density of 724.67） was significantly lower than that of COC group and COC＋GC group （P0.05）. Detection of glucose-6-phosphate dehydrogenase （G6PDH） activity showed that there was no significant difference in BCB positive oocyte rate between DO ＋MGC group （88.26% ） and COC group （92.75%） or between DO＋MGC group and DO group （82.86% ）, however, BCB positive oocyte rate of DO group was significantly lower than that of COC group （P0.05）. Furthermore, the cleavage rate and blastocyst rate of activated mature oocytes derived from DO ＋MGC group （94.98% and 43.67% , respectively） were significantly higher than those from DO group （52.54% and 8.97%, respectively） （P0.05）, and were not significantly different compared with those from COC group （97.11% and 38.30%, respectively）. In addition, the cleavage rate of fertilized oocytes derived from DO＋MGC group （72.65%） showed no significant difference compared with that from DO group （63.59%）, but the blastocyst rate of DO＋MGC group was significantly higher than that of DO group （9.88%） （P0.05）. [Conclusion] MGC coculture can significantly improve the in vitro cytoplasmic maturation quality of denuded porcine oocytes, thereby enhancing the subsequent developmental potential.

Establishment on Isolated Microspore Culture Optimized System for Restorer of New Cytoplasmic Male Sterile (NER) of Brassica napus L.

[Objective]The aim was to establish optimized system for NER isolated microspore culture of Brassica napus L.. [Method]Twenty varieties of NER of Brassica napus were grown under uncontrolled temperature and light conditions,and their isolated microspore from anthers were used as explants in vitro culture. The influencing factors of microspore culture were preliminarily studied. [Result]The difference of genotypes was important influencing factors to embryoid yield. The embryoid yield increased by supplementing with 6-BA and NAA,culturing in solid-liquid double layer medium with activated charcoal; The difference was not significant of embryoid yield between culturing in medium supplemented with colchicines and the CK. The rates of cotyledonous embryoids directly developed into normal plantlets increased through enriching with 0.1-0.2 mg/L NAA and being treated on slim illumination two days before being inducted into normal plantlets. [Conclusion]The technical system of microspore culture of restorer of new cytoplasmic male sterile （NER） was established,which and lays a foundation for accelerating genotype purification of NER introgressive line.

Cytological Mechanism of Cytoplasmic Inheritance in Pinus tabulaeformis: Ⅱ. Transmission of Male and Female Organelles During Fertilization and Proembryo Development

In an earlier report the ultrastructure and nucleoid organelles of male gamete in Pinus tabulaeformis Carr. have been described. Presently, the ultrastructure of the cytoplasm of the egg cell and pollen tube—immediately before fertilization and during cytoplasmic transmission of male gametophyte—has been described for the same species. The fate of parental plastids and mitochondria in the proembryo has also been followed. The mature egg cell contains a large amount of mitochondria, but seems to lack normal plastids. Most plastids have transformed into large inclusions. Apart from the large inclusions, there are abundant small inclusions and other organelles in the egg cell. During fertilization, pollen tube penetrates into the egg cell at the micropylar end and thereafter the contents are released. Plastid and mitochondrion of male origin are lacking near the fusing sperm_egg nuclei. The second sperm nucleus—not involved in karyogamy—remains at a site near the receptive vacuole. This nucleus is surrounded by large amount of male cytoplasm containing mixed organelles from the sperm cell, tube cell, and egg cell. At the free nuclear proembryo stage, organelles of male and female origin are visible in the perinucleus_cytoplasmic zone. Most of the mitochondria have the same morphological features as those in the egg cell. Some of the mitochondria appear to have originated from the sperm and tube cells. Plastids are most likely of male gametophyte origin because they have similar appearance as those of the sperm and tube cell. Large inclusions in the egg cell become vacuole_like. Paternal plastids have been incorporated into the neocytoplasm of the proembryo. In the cellular proembryo, maternal mitochondria are more abundant. Plastids resembling those of the sperm and tube cell are still present. These cytological results clearly show that in P. tabulaeformis, plastids are inherited paternally and mitochondria bipaternally. The cytological mechanism of plastid and mitochondrion inheritance in gymnosperm is discussed.