Identification of candidate genes for early-maturity traits by combining BSA-seq and QTL mapping in upland cotton(Gossypium hirsutum L.)

Cotton breeding for the development of early-maturing varieties is an effective way to improve multiple cropping indexes and alleviate the conflict between grains and cotton in the cultivated fields in China.In the present study,we aimed to identify upland cotton quantitative trait loci(QTLs)and candidate genes related to early-maturity traits,including whole growth period(WGP),flowering timing(FT),node of the first fruiting branch(NFFB),height of the node of the first fruiting branch(HNFFB),and plant height(PH).An early-maturing variety,CCRI50,and a latematuring variety,Guoxinmian 11,were crossed to obtain biparental populations.These populations were used to map QTLs for the early-maturity traits for two years(2020 and 2021).With BSA-seq analysis based on the data of population 2020,the candidate regions related to early maturity were found to be located on chromosome D03.We then developed 22 polymorphic insertions or deletions(InDel)markers to further narrow down the candidate regions,resulting in the detection of five and four QTLs in the 2020 and 2021 populations,respectively.According to the results of QTL mapping,two candidate regions(InDel_G286-InDel_G144 and InDel_G24-InDel_G43)were detected.In these regions,three genes(GH_D03G0451,GH_D03G0649,and GH_D03G1180)have nonsynonymous mutations in their exons and one gene(GH_D03G0450)has SNP variations in the upstream sequence between CCRI50 and Guoxinmian 11.These four genes also showed dominant expression in the floral organs.The expression levels of GH_D03G0451,GH_D03G0649 and GH_D03G1180 were significantly higher in CCRI50 than in Guoxinmian 11 during the bud differentiation stages,while GH_D03G0450 showed the opposite trend.Further functional verification of GH_D03G0451 indicated that the GH_D03G0451-silenced plants showed a delay in the flowering time.The results suggest that these are the candidate genes for cotton early maturity,and they may be used for breeding early-maturity cotton varieties.

A novel transcription factor FnMYB4 regulates pigments metabolism of yellow leaf mutants in Fragaria nilgerrensis

The strawberry species Fragaria nilgerrensis Schlechtendal ex J.Gay,renowned for its distinctive white,fragrant peach-like fruits and strong disease resistance,is an exceptional research material.In a previous study,an ethyl methane sulfonate(EMS)mutant library was established for this species,resulting in various yellow leaf mutants.Leaf yellowing materials are not only the ideal materials for basic studies on photosynthesis mechanism,chloroplast development,and molecular regulation of various pigments,but also have important utilization value in ornamental plants breeding.The present study focused on four distinct yellow leaf mutants:mottled yellow leaf(MO),yellow green leaf(YG),light green leaf(LG),and buddha light leaf(BU).The results revealed that the flavonoid content and carotenoid-to-chlorophyll ratio exhibited a significant increase among these mutants,while experiencing a significant decrease in chlorophyll and carotenoid contents compared to the wild type(WT).To clarify the regulatory mechanisms and network relationships underlying these mutants,the RNA-seq and weighted gene coexpression network(WGCNA)analyses were employed.The results showed flavonoid metabolism pathway was enriched both in MO and YG mutants,while the chlorophyll biosynthesis pathway and carotenoid degradation pathway were only enriched in MO and YG mutants,respectively.Subsequently,key structural genes and transcription factors were identified on metabolic pathways of three pigments through correlation analyses and quantitative experiments.Furthermore,a R2R3-MYB transcription factor,FnMYB4,was confirmed to be positively correlated with flavonoid synthesis through transient overexpression,virus-induced gene silencing(VIGS),and RNA interference(RNAi),accompanying by reoccurrence and attenuation of mutant phenotype.Finally,dual-luciferase(LUC)and yeast one-hybrid assays confirmed the binding of FnMYB4 to the FnFLS and FnF3H promoters,indicating that FnMYB4 positively regulates flavonoid synthesis.In addition,correlation analyses suggested that FnMYB4 also might be involved in chlorophyll and carotenoid metabolisms.These findings demonstrated the pivotal regulatory role of FnMYB4 in strawberry leaf coloration.

Detection of ARV-Resistant Mutants in HIV-1-Infected Individuals in a Context of Systematic Switching to an Association Based on Dolutegravir in Abidjan, Côte d’Ivoire

The emergence of antiretroviral resistance mutations represents a major threat to the achievement of national and global goals for the elimination of HIV-1 infection. The global strategy in 2019 in Cte d'Ivoire is a new national policy for the management of people living with HIV with the administration of dolutegravir (DTG)-based fixed-dose combination. The aim of our study was to evaluate HIV-1 resistance to antiretrovirals (ARVs) in infected adult subjects in Cte d’Ivoire in the context of a systematic switch to a DTG-based combination. Between February 2022 and October 2023, a cross-sectional survey with random sampling was conducted in 06 services caring for people living with HIV. A total of 139 participants were included in the study. Adults with a viral load ≥ 1000 copies/mL were tested for HIV-1 ARV resistance mutations. Molecular analyses were performed using protocol of ANRS-MIE (National Agency for Research on AIDS and emerging infectious diseases). The interpretation is performed by HIVGRAD (https://www.hiv-grade.de/cms/grade/). The frequencies of HIV-1 resistance to non-nucleotide reverse transcriptase inhibitors (NNRTIs), nucleotide reverse transcriptase inhibitors (NRTIs), integrase inhibitors (IINTs) and protease inhibitors (PIs) were 82%, 73%, 19% and 11% respectively. The main mutations observed in the different classes were K103N (45%), M184V (64%), E157Q (19%) and L10V/M46I/A71V/I54V (6%) respectively. This study reveals the emergence of resistance to DTG-based fixed-dose combinations, favored by high rates of resistance to NRTIs and NNRTIs. This finding underlines the need for enhanced viral load monitoring and HIV-1 genotyping tests to guide the choice of NRTIs for combination therapy. In addition, monitoring for mutations to second-generation NRTIs is essential, given the scale-up of DTG-based regimens currently underway in Cte d’Ivoire.

An efficient method for constructing a random insertional mutant library for forward genetics in Nannochloropsis oceanica

Insertional mutation,phenotypic evaluation,and mutated gene cloning are widely used to clone genes from scratch.Exogenous genes can be integrated into the genome during non-homologous end joining(NHEJ)of the double-strand breaks of DNA,causing insertional mutation.The random insertional mutant library constructed using this method has become a method of forward genetics for gene cloning.However,the establishment of a random insertional mutant library requires a high transformation efficiency of exogenous genes.Many microalgal species show a low transformation efficiency,making constructing random insertional mutant libraries difficult.In this study,we established a highly efficient transformation method for constructing a random insertional mutant library of Nannochloropsis oceanica,and tentatively tried to isolate its genes to prove the feasibility of the method.A gene that may control the growth rate and cell size was identified.This method will facilitate the genetic studies of N.oceanica,which should also be a reference for other microalgal species.

Roles of Mutant TP53 Gene in Cancer Development and Progression

TP53 is a tumor suppressor gene that is mutated in most cancer types and has been extensively studied in cancer research.p53 plays a critical role in regulating the expression of target genes and is involved in key processes such as apoptosis,cell cycle regulation,and genomic stability,earning it the title“guardian of the genome.”Numerous studies have demonstrated p53’s influence on and regulation of autophagy,ferroptosis,the tumor microenvironment,and cell metabolism,all of which contribute to tumor suppression.Alterations in p53,specifically mutant p53(mutp53),not only impair its tumor-suppressing functions but also enhance oncogenic characteristics.Recent data indicate that mutp53 is strongly associated with poor prognosis and advanced cancers,making it an ideal target for the development of novel cancer therapies.This review summarizes the post-translational modifications of p53,the mechanisms of mutp53 accumulation,and its gain-of-function,based on previous findings.Additionally,this review discusses its impact on metabolic homeostasis,ferroptosis,genomic instability,the tumor microenvironment,and cancer stem cells,and highlights recent advancements in mutp53 research.

Yield Stability of Direct-sowing and Early-maturing Excellent Brassica napus (Attar No.8)

[Objective] The aim was to analyze yield stability of attar No.8, which is direct sowing and early-maturing. [Method] Analysis was made on yielding ability, stability and adaptability of attar No.8 in Yunnan Province during 2006-2009 with DPS software and stability assessment was conducted on yield and major economic characters of the variety in different periods and experimental plots. [Result] Average yield of attar No.8 during three years in 24 plots achieved 3 379.17 kg/hm2; it grew tightly and is featured with early matureness. In addition, grain density, pod number and annul production differed little in different years and the variety in eight ecological conditions all showed high production potential, yielding ability and stability, which proved that attar No.8 is suitable to be grown in Yunnan and southern early-maturing areas in China. [Conclusion] The research provided references for selection and breeding of rapeseeds＇ variety which enjoys high and stable yield.

Variations in chlorophyll content, stomatal conductance, and photosynthesis in Setaria EMS mutants

Chlorophyll (Chl) content,especially Chl b content,and stomatal conductance (G_s) are the key factors affecting the net photosynthetic rate (P_n).Setaria italica,a diploid C_4 panicoid species with a simple genome and high transformation efficiency,has been widely accepted as a model in photosynthesis and drought-tolerance research.The current study characterized Chl content,G_s,and P_n of 48 Setaria mutants induced by ethyl methanesulfonate.A total of 24,34,and 35 mutants had significant variations in Chl content,G_s,and P_n,respectively.Correlation analysis showed a positive correlation between increased G_s and increased P_n,and a weak correlation between decreased Chl b content and decreased P_n was also found.Remarkably,two mutants behaved with significantly decreased Chl b content but increased P_n compared to Yugu 1.Seven mutants behaved with significantly decreased G_s but did not decrease P_(n )compared to Yugu 1.The current study thus identified various genetic lines,further exploration of which would be beneficial to elucidate the relationship between Chl content,G_s,and P_n and the mechanism underlying why C_4 species are efficient at photosynthesis and water saving.

Influence of the Number of Reserved Fruit on Fruit-bearing Branch of Early-maturing Citrus unshiu Marc. on Fruit Growth and Development

In order to provide theoretical references for early-maturing Citrus unshiu Marc. in terms of fruit thinning, timely fertilization and harvesting, as wel as for for-mulation of of Early-maturing Citrus unshiu Marc. Cultivation Technique Standards in Guizhou Province, the dynamic changes of fruit longitudinal diameter and trans-verse diameter of 11-year-old and 12-year-old early maturing Citrus unshiu Marc. were tested in two consecutive years and the effects of single-fruit bearing branch on fruit growth and development were compared. The results showed annual varia-tion of longitudinal and transverse diameter of the fruit in Guizhou grew faster at the beginning and then slowed down gradual y, maintaining a slow-growth period after October; the growth curve final y was flat in mature stage. General y, the growth and development stage of fruit showed four growth peaks, showing an in-creasing S-shape curve and fruit net increase also had four growth peaks with a double S-shape curve. Specifical y, longitudinal diameters growth alternated with transverse diameter growth and the former proved much stronger. In terms of ef-fects of single-fruit bearing branch on longitudinal and transverse diameter growth, the number of reserved fruits had direct effects on the growth, net increase of weight, time of growth peak, fluctuation and the number of growth peak. The growth and development, and quality of fruits in the treatments with two or three fruits re-served on single fruit-bearing branch proved better compared with the control with four left fruits. It suggested that early June and early July are the best time for fer-tilization of early-maturing Citrus unshiu Marc. in Guizhou Pprovince and would have better effect if combined with foliage spraying. The suitable harvest time is in the end of October. The preferred number of reserved fruit on single fruit-bearing branch is 2-3. In actual production, the implementation of flower and fruit thinning should also takes tree age, balance of tree vigor, strength of fruit bearing branch, alternate bearing phenomenon and other factors into consideration.

Construction of EMS-Induced Peanut Mutant Libraries and Identification of Pod-Related Traits Mutant Lines

Peanut(Arachis hypogaea L.)is an oil and economic crop of vital importance,and peanut pod is the key organ influencing the yield and processing quality.Hence,the Pod-related traits(PRTs)are considered as important agronomic traits in peanut breeding.To broaden the variability of PRTs in current peanut germplasms,three elite peanut cultivars were used to construct Ethyl methane sulfonate(EMS)-induced mutant libraries in this study.The optimal EMS treatment conditions for the three peanut varieties were determined.It was found that the median lethal dose(LD50)of EMS treatment varied greatly among different genotypes.Finally,the EMS-induced peanut mutant libraries were constructed and a total of 124 mutant lines for PRTs were identified and evaluated.Furthermore,“M-8070”,one of the mutant lines for pod constriction,was re-sequenced via high-throughput sequencing technology.The genome-wide variations between“M-8070”and its wild parent“Fuhua 8”(FH 8)were detected.2994 EMS-induced single nucleotide polymorphisms(SNPs)and 1188 insertion-deletions(InDels)between“M-8070”and its wild parent were identified.The predominant SNP mutation type was C/G to T/A transitions,while the predominant InDel mutation type was“1-bp”.We analyzed the distribution of identified mutations and annotated their functions.Most of the mutations(91.68%of the SNPs and 77.69%of the InDels)were located in the intergenic region.72 SNPs were identified in the exonic region,leading to 27 synonymous,43 nonsynonymous and 2 stop-gain variation for gene structure.13 Indels were identified in the exonic region,leading to 4 frame-shift,8 non-frame-shift and 1 stop-gain variations of genes.These mutations may lead to the phenotypic variation of“M-8070”.Our study provided valuable resources for peanut improvement and functional genomic research.

Transcriptome Analysis of Molecular Mechanisms Underlying Phenotypic Variation in Phaseolus vulgaris Mutant‘nts’

The phenotype of a common bean plant is often closely related to its yield,and the yield of plants with reduced height or poor stem development during growth is low.Mutants serve as an essential gene resource for common bean breeding genetic research.Although model plants and crops are studied to comprehend the molecular mechanisms and genetic basis of plant phenotypes,the molecular mechanism of phenotypic variation in common beans remains underexplored.We here used the mutant‘nts’as material for transcriptome sequencing analysis.This mutant was obtained through 60Co-γirradiation from the common bean variety‘A18’.Differentially expressed genes were mainly enriched in GO functional entries such as cell wall organization,auxin response and transcription factor activity.Metabolic pathways significantly enriched in KEGG analysis included plant hormone signal transduction pathways,phenylpropanoid biosynthesis pathways,and fructose and mannose metabolic pathways.AUX1(Phvul.001G241500),the gene responsible for auxin transport,may be the key gene for auxin content inhibition.In the plant hormone signal transduction pathway,AUX1 expression was downregulated and auxin transport across the membrane was blocked,resulting in stunted growth of the mutant‘nts’.The results provide important clues for revealing the molecular mechanism of‘nts’phenotype regulation in bean mutants and offer basic materials for breeding beneficial phenotypes of bean varieties.

Mapping and Functional Analysis of LE Gene in a Lethal Etiolated Rice Mutant at Seedling Stage

An EMS(ethy methanesulfonate)-induced lethal etiolated(le)mutant obtained from the rice variety Zhongjian 100 was characterized by lethal etiolated phenotypes,with significantly reduced levels of chlorophyll a,chlorophyll b,total chlorophyll,and carotenoids.Additionally,the mutant displayed a significantly decreased number of chloroplast grana,along with irregular and less-stacked grana lamellae.The le mutant showed markedly diminished root length,root surface area,and root volume compared with the wild type.It also exhibited significantly lower catalase activity and total protein content,while peroxidase activity was significantly higher.Using the map-based cloning method,we successfully mapped the LE gene to a 48-kb interval between markers RM16107 and RM16110 on rice chromosome 3.A mutation(from T to C)was identified at nucleotide position 692 bp of LOC_Os03g59640(ChlD),resulting in a change from leucine to proline.By crossing HM133(a pale green mutant with a single-base substitution of A for G in exon 10 of ChlD subunit)with a heterozygous line of le(LEle),we obtained two plant lines heterozygous at both the LE and HM133 loci.Among 15 transgenic plants,3 complementation lines displayed normal leaf color with significantly higher total chlorophyll,chlorophyll a,and chlorophyll b contents.The mutation in le led to a lethal etiolated phenotype,which has not been observed in other ChlD mutants.The mutation in the AAA+domain of ChlD disrupted the interaction of ChlDle with ChlI as demonstrated by a yeast two-hybrid assay,leading to the loss of ChlD function and hindering chlorophyll synthesis and chloroplast development.Consequently,this disruption is responsible for the lethal etiolated phenotype in the mutant.

西瓜果实网条突变体的转录组分析

以条纹自交系西瓜(TD)和网条突变体西瓜(WT)为材料,通过分离群体的条纹性状比例探索西瓜的条性状遗传基础;以转录组测序及表达差异分析为基础,探索网条突变体西瓜在不同发育阶段的表达谱变化,筛选出条纹自交系与网条突变体西瓜之间表达差异显著的基因57个,主要富集于光合作用天线蛋白通路、生物碱生物合成通路、氨基酸代谢通路等7种代谢途径;通过加权基因共表达网络对核心基因的进一步分析,初步挖掘出部分网条突变体西瓜中有关生物胁迫抗性的基因变化情况,进一步筛选确定西瓜深绿条纹基因(ClGS,watermelon dark-green stripe)、类半胱氨酸蛋白酶抑制剂5编码基因、UDP-葡萄糖基转移酶编码基因、类长春碱合成酶编码基因、叶绿素a-b结合蛋白编码基因可能是决定西瓜网条果皮性状的核心基因;qRT-PCR验证结果表明,条纹自交系西瓜与网条突变体西瓜在基因表达上存在明显差异。本研究为西瓜不同果皮条纹育种提供了新的种质资源,也为阐明西瓜不同果皮条纹性状形成的分子调控机制提供了一定的理论依据。

靶向敲除棉花GhAGL16高效sgRNA的筛选

【目的】AGL16是棉花MADS-box基因家族中一个重要的转录负调控因子,在抵御干旱和盐胁迫过程中发挥着重要作用。利用病毒介导的基因编辑技术(virus-induced gene editing,VIGE)筛选靶向敲除棉花GhAGL16的sgRNAs,并验证这些sgRNAs的特异性,为定向创制棉花agl16突变体奠定重要基础。【方法】根据在棉花YZ-1中A亚组和D亚组上实际克隆GhAGL16的基因组序列,预测了3个能靶向敲除GhAGL16的sgRNAs;基于棉花叶皱缩病毒(cotton leaf crumple virus,CLCrV)介导的VIGE系统,构建3个CLCrV-AtU6-26::GhAGL16-sgRNAs表达载体;利用qPCR检测Cas9超表达(Cas9 over-expression,Cas9-OE)植株中Cas9的表达量,确定Cas9是否稳定遗传表达;将3个CLCrV-AtU6-26::GhAGL16-sgRNAs表达载体分别转化Cas9-OE棉花子叶,并通过PCR/RE方法检测3个靶点的突变情况;利用生物信息学方法分析3个GhAGL16-sgRNAs的二级结构;对突变植株进行Hi-TOM高通量测序,明确基因编辑效率。同时,对转化GhAGL16-sgRNA2的突变植株进行脱靶率鉴定,检测基因编辑的特异性。【结果】成功构建了3个能同时靶向敲除GhAGL16-A亚组和GhAGL16-D亚组序列的sgRNAs。对不同Cas9-OE植株中Cas9表达量检测结果表明,Cas9在不同Cas9-OE棉株中稳定表达。用3个CLCrV-AtU6-26::GhAGL16-sgRNAs表达载体转化Cas9-OE棉花子叶,PCR/RE突变检测结果表明,GhAGL16-sgRNA2可有效用于GhAGL16的靶向敲除,在棉花A亚组和D亚组靶位点上出现了不同碱基缺失的突变类型,而GhAGL16-sgRNA1和GhAGL16-sgRNA3是2个无效sgRNA。3个GhAGL16-sgRNAs的二级结构分析结果表明,GhAGL16-sgRNA1和GhAGL16-sgRNA3可能存在引导序列容易与其他序列发生配对并且不易解链的现象,干扰了引导序列对目标位点的识别,导致sgRNA无效。进一步量化GhAGL16-sgRNA2对GhAGL16的编辑效率,对每一株转化CLCrV-AtU6-26::GhAGL16-sgRNA2的Cas9-OE植株突变检测结果表明,在转化的9株Cas9-OE植株中有6株发生了突变,突变效率为66.67%。此外,Hi-TOM高通量测序结果表明,GhAGL16-sgRNA2对GhAGL16的编辑效率为13.69%—54.42%。脱靶率鉴定结果表明,预测的4个潜在脱靶位点都没有发现脱靶现象,说明GhAGL16-sgRNA2不仅具有高效的基因编辑效率,而且具有专一的基因编辑特异性。【结论】利用CLCrV介导的VIGE系统转化Cas9-OE棉花筛选获得1个能高效敲除GhAGL16的sgRNA,为定向创制棉花agl16突变体提供了理想的sgRNA。

玉米矮秆突变体20F421的表型鉴定及遗传分析

矮秆资源是农作物矮化育种的物质基础,发掘矮秆基因资源对培育矮秆新品种具有重要作用。为了明确^(252)CF裂变快中子辐射诱变玉米自交系KWS49筛选得到的矮秆突变体20F421的遗传特性和矮化机理,以20F421为材料分别与玉米自交系PH6WC、B73、Mo17及KWS49杂交构建F_(1)和F_(2)分离群体,分析矮秆性状的遗传模式,并以(20F421/B73)F_(2)为定位群体,采用混池转录组测序(bulked segregant RNA-seq,BSR-seq)方法初步定位突变基因。结果表明,与KWS49相比,20F421的植株高度为95.2 cm,降低47.89%;穗位高度为23.9 cm,降低64.54%;茎秆节间长度显著缩短、叶片较直立密生,自交结实良好。遗传分析表明,F2分离群体野生型(高秆)与突变型(矮秆)植株性状分离比例符合3∶1,表明该突变体受单个核隐性基因控制;BSR-seq结果将突变基因定位在1号染色体177~255 Mb之间。通过与B73参考基因组进行比对发现,该区间内含有矮秆基因Br2,将20F421与br2突变体杂交进行等位性检测,F_(1)和F_(2)的株高均没有发生性状分离,表现为突变体20F421和br2表型,推测20F421的矮秆突变基因为矮秆基因Br2的等位突变。研究结果为进一步精细定位、克隆突变基因奠定基础,也为解析玉米矮化机理和培育矮秆玉米新品种提供重要基因资源和理论支撑。

抗枯萎病宝岛蕉早花突变体的筛选与鉴定

【目的】筛选鉴定抗枯萎病香蕉品种宝岛蕉的早花突变体,为进一步利用突变体株系选育适应性更广的香蕉新品种提供参考依据。【方法】利用多代组织培养繁育结合田间种植对比试验,依据生育期短、抗病性强、株高矮、株产高和遗传稳定筛选目标,从宝岛蕉早花突变后代中筛选优良株系,以宝岛蕉为对照,按照香蕉种质资源描述规范对优良突变株系的形态特征和生物学特性等进行观察鉴定,测定分析植株球茎生长点部位碳、氮营养累积以及成花相关激素吲哚乙酸(IAA)、玉米素(ZR)、赤霉素(GA)和脱落酸(ABA)含量差异,通过ISSR分子标记检测突变体及其野生型之间的遗传变异和亲缘关系。以宝岛蕉和主栽品种(桂蕉6号及巴西蕉)为对照,利用伤根浸菌法进行苗期抗病性鉴定,并在广西和海南开展区域种植试验及田间抗病性鉴定。【结果】筛选获得优良突变株系0523(简称0523),其新植蕉平均株高280.0 cm,假茎基围80.5 cm、中围60.0 cm,较宝岛蕉分别减小14.0%、5.8%和11.8%,新抽总叶片数26~29片;果实营养品质和单株产量与宝岛蕉无显著差异(P>0.05,下同)。0523球茎生长点碳氮比较宝岛蕉显著提高19.0%(P<0.05,下同),GA和IAA含量显著降低24.8%和22.6%,ABA和ZR含量与宝岛蕉相比略有增加,差异不显著。抗病性苗期鉴定结果显示0523为中抗且偏强,抗性水平与宝岛蕉相比略有提高,桂蕉6号为高感。突变株系0523与宝岛蕉的多态性比率为8.8%,遗传相似系数为0.95,二者存在差异。0523在广西和海南各试验点第1、2造蕉平均发病率分别为4.5%和3.5%,较主栽品种降低87.8%和94.1%,各试验点发病率均显著低于主栽品种。0523第1、2造蕉的生育期较宝岛蕉显著缩短,与主栽品种无显著差异。【结论】突变株系0523具有抗枯萎病、早熟、矮化、适应性广等优良性状,可用来培育大面积推广的香蕉新品种,或作为亲本育种材料应用于香蕉育种研究。

代谢组与转录组联合解析赤皮青冈叶片黄化变异机制

为揭示赤皮青冈叶色黄化变异机制,该研究以赤皮青冈叶色变异植株和正常植株的叶片为试验材料,采用超高效液相色谱串联质谱法和高通量RNA测序技术分别进行代谢组和转录组分析。结果表明:(1)代谢组在正离子(POS)、负离子(NEG)模式下分别检测出正常植株和突变体之间存在257个和357个显著差异代谢物(SCMs),其中槲皮素、白矢车菊素、杨梅素等多种黄酮类化合物及其糖苷衍生物(吡喃酮啡肽A、异鼠李素3-葡糖苷酸等)在突变体中显著上调,而叶绿素a、叶绿素b、类胡萝卜素等色素含量则显著下降。(2)转录组测序检测出4146个差异表达基因(DEGs),其中1711个基因上调表达,2435个基因下调表达。(3)KEGG富集分析表明,SCMs和DEGs显著富集到光合作用、卟啉与叶绿素代谢、类黄酮生物合成等途径。综上表明,突变体叶色黄化可能是受到叶绿素合成受阻、叶绿体发育异常及黄酮物质合成增加等因素的综合影响。此外,MYB和bHLH家族基因在突变体中显著上调,证实该两类转录因子参与调控类黄酮生物合成。该研究结果为植物黄化突变的分子机制研究提供了新的见解,也为叶色功能基因挖掘与园林植物育种工作提供了参考。

大白菜蜡质缺失突变体YW71的遗传及序列变异分析

以大白菜蜡质突变体YW71为对象,研究其亮绿无蜡粉性状的遗传规律和调控基因。通过遗传分析,表明YW71中的亮绿无蜡粉性状由隐性单基因控制。通过等位基因互补实验证明YW71的亮绿性状是BrWAX2等位突变引起的。序列分析表明,在YW71中BrWAX2基因在第3个外显子末端发生了39 bp的缺失,进而引起转录水平的可变剪切和翻译水平的提前终止。表达模式分析表明,BrWAX2基因在YW71茎和叶中表达量显著下降。此外,研究针对39 bp的变异开发并验证了共显性标记BrWAX2-In Del1。研究结果丰富了白菜类蔬菜蜡质突变遗传资源,将为亮绿无蜡粉品种的分子育种提供理论指导和技术支持。

一株拟南芥宽叶形突变体atscamp的分离鉴定

叶片是主要的光合作用器官,选育利于光合作用的叶片形态已成为重要的育种目标。atscamp是从拟南芥(Arabidopsis thaliana)突变体库(约6000株系)中筛选获得的1株叶片宽大突变体。Tail-PCR分析该突变体为AT1G11180位点的插入,该基因位点编码1个分泌载体膜蛋白(SCAMP)。RT-PCR检测显示,该基因转录表达水平基本为零。进一步研究发现,该突变体叶片的宽度和叶面积极显著大于野生型植株(P<0.01),但是冠幅基本保持不变;同时atscamp突变体叶绿素含量增加,叶绿素最大荧光、PSⅡ潜在光化学效率显著增加(P<0.05);相应地,突变体植株蒸腾系数(Tr)、净光合速率(Pn)和叶片水分利用效率(WUE)显著增加(P<0.05)。拟南芥AT1G11180基因的时空特异性表达分析显示,该基因仅在叶片中高表达,在其他器官中表达量很低;且随着植物发育成熟,该基因表达量逐渐增加。研究结果表明AtSCAMP基因在叶形发育中发挥着重要作用。

基于变异的正则表达式反例测试串生成算法

正则表达式在计算机科学的许多领域具有广泛应用.然而,由于正则表达式语法比较复杂,并且允许使用大量元字符,导致开发人员在定义和使用时容易出错.测试是保证正则表达式语义正确性的实用和有效手段,常用的方法是根据被测表达式生成一些字符串,并检查它们是否符合预期.现有的测试数据生成大多只关注正例串,而研究表明,实际开发中存在的错误大部分在于定义的语言比预期语言小,这类错误只能通过反例串才能发现.研究基于变异的正则表达式反例测试串生成.首先通过变异向被测表达式中注入缺陷得到一组变异体,然后在被测表达式所定义语言的补集中选取反例字符串揭示相应变异体所模拟的错误.为了能够模拟复杂缺陷类型,以及避免出现变异体特化而无法获得反例串的问题,引入二阶变异机制.同时采取冗余变异体消除、变异算子选择等优化技术对变异体进行约简,从而控制最终生成的测试集规模.实验结果表明,与已有工具相比,所提算法生成的反例测试串规模适中,并且具有较强的揭示错误能力.

苜蓿抗寒突变体生理生化及性状指标分析

为选育在黑龙江省可越冬的理想紫花苜蓿品种。以‘龙牧806’零磁空间诱变处理后二代群体中获得的lm1609、lm1625、lm1704三个抗寒突变体为实验材料,通过测定发芽指标、越冬率、产量特性指标及根系生理生化指标,进行抗寒性能鉴定。3个突变体发芽指数与对照差异不显著;发芽势、简化活力指数lm1609显著高于lm1625、lm1704,分别为82.05%、469.82(P<0.05);3个突变体的越冬率都显著低于对照组;突变体和对照6月6日起始株高差异不显著,随着植株生长,7月6日和8月6日lm1609、lm1704显著高于lm1625和对照;突变体lm1609、lm1625、lm1704三者鲜草产量差异不显著,但显著高于对照(P<0.05);三个苜蓿突变体植株根系过氧化物酶、超氧化物歧化酶均显著高于对照,并且突变体lm1609 POD、SOD最高,分别达到304.85 OD470/(min·g)、208.41 U/g(P<0.05)。lm1609突变体具有较高的抗寒特性及产量特性,是为理想的抗寒突变体材料。