玉米籽粒突变体dek48的表型鉴定与基因定位

籽粒是玉米的主要营养储存器官,也是禾本科植物种子发育研究的模式器官。本研究对玉米自交系郑58进行甲基磺酸乙酯(EMS)诱变,获得一个稳定遗传的籽粒缺陷突变体,命名为defective kernel 48 (dek48)。该突变体籽粒皱缩扁小,百粒重显著降低,胚和胚乳发育严重缺陷,不能成苗。在玉米授粉后12 d即可观察到明显的发育缺陷,表明该突变发生在籽粒发育的早期阶段。扫描电镜观察发现dek48与野生型相比淀粉粒显著变小。石蜡切片显微观察发现dek48淀粉胚乳填充不饱满,糊粉层细胞发育不规则。遗传学分析表明,该突变性状受隐性单基因控制。进一步构建F2遗传定位群体,将该突变体基因精细定位于3号染色体7.39 Mb～7.52 Mb之间。生物信息学分析发现该区间内有6个开放阅读框,暂未发现与籽粒发育有关的已知基因,后续将通过测序和基因表达分析进一步确定候选基因。     

玉米籽粒突变体smk7的表型分析和基因定位

In this study, a stable small kernel mutant, named small kernel 7 (smk7), was isolated from ethylmethane sulfonate (EMS) mutagenesis of maize inbred line B73. Compared with wild type, the smk7 mutants showed smaller kernel size, defective embryo and endosperm development and a significant decrease in 100-kernel weight. The smk7 kernels showed a low level of germination rate at 10% and cannot grow into normal plants. No significant changes were detected in protein, starch and oil content between mature wild type and smk7 kernels, but the starch grains became significantly smaller and irregular in smk7 kernels compared with wild type. The smk7 kernels could be clearly distinguished from the wild type as early as 12 days after pollination (DAP), on the basis of their smaller and emptier phenotype. Microscopic inspection of the paraffin sections revealed that the development of embryo and endosperm were delayed, and the cell wall in growth in basal endosperm transfer layers (BETL) were arrested in smk7 compared with wild type. The F₂ populations with multiple backgrounds were constructed by crossing heterozygous plants (+/smk7) with several other inbred lines. Genetic analysis showed that the mutant phenotype was controlled by a single recessive gene. Based on genotyping by target sequencing (GBTS) strategy, the SMK7 was initially mapped on the short arm of chromosome 2. The fine mapping results suggested that SMK7 was located between markers RM1433917 and RM1535316, with a physical distance of 120 kb. There were eight protein-coding genes in this region. This study laid a foundation for further genes cloning and research of the SMK7 function in regulating maize kernel development.     

玉米F2群体轴部相关性状的数量遗传分析

本研究采用遗传模型预测的方法，探究玉米轴部性状的遗传机理，以期为育种家指明育种方向，提高育种效率。以FBHJ和MBUB两亲本组配的含390个家系的F₂群体为材料，利用植物数量分离分析软件包中的F₂单世代分析方法，对其穗轴长、粒长、穗轴粗、粒宽、穗轴重、百粒重、粒厚等7个轴部相关性状进行数量遗传分析、相关分析以及回归分析。结果表明，穗轴粗、粒长、粒宽3个性状最适遗传模型为A-0，即无主基因遗传；穗轴重和粒厚受2对主基因控制，主基因存在加性-显性-上位性，符合B-1模型；穗轴长和百粒重由1对主效基因控制，最适模型分别为A-1、A-3，各模型主基因遗传率介于28.37%～59.11%，其中百粒重性状的遗传率最大。各性状均与百粒重呈极显著正相关。百粒重(Y)对其他农艺性状的回归方程为Y=-57.126+0.176X₁+3.069X₂+2.890X₃+5.108X₄，拟合度为0.723。通径分析中粒宽的间接作用和粒长的直接作用表现最大。这一研究结果为玉米轴部性状的遗...     

玉米穗发芽突变体vp-like8的遗传分析及突变基因鉴定

玉米突变体vp-like8具有明显的穗发芽性状且能稳定遗传,遗传分析表明该突变性状受隐性单基因控制。用vp-like8与自交系郑58杂交构建F2遗传定位群体,利用BSR-Seq方法,将基因初定在玉米第3染色体160.4 Mb～165.6Mb区间内。参考玉米基因组信息,发现已报道的穗发芽基因Vp1位于此定位区间内。分别利用vp1、vp-like8的杂合突变体进行等位测验,发现杂交后代中正常与穗发芽籽粒符合3∶1遗传分离比。经序列分析,发现vp-like8突变体中Vp1基因在第2内含子有343 bp碱基的缺失,且第3内含子有222 bp重复序列的插入,而已报道的vp1突变体只在第2个内含子有343 bp碱基的缺失。通过实时定量PCR检测发现,与正常籽粒相比, vp-like8与Vp1突变籽粒中Vp1基因的转录水平均明显降低。以上证据表明, vp-like8是一个新的Vp1基因等位突变体。     

水稻甜质胚乳突变体m5788的鉴定及基因定位

胚乳发育是种子形成的关键,其决定水稻的外观品质和食味品质。m5788是从粳稻品种中花11的组织培养后代中发现的甜质胚乳突变体,其籽粒皱缩,千粒重与穗粒数均显著降低,淀粉合成受阻,可溶性糖含量显著增加。通过对m5788与IRAT129杂交产生的F₂代群体分析表明,甜质胚乳性状受1对隐性核基因控制。对569个F₂隐性极端单株进行连锁分析和定位,将目的基因定位在8号染色体长臂端Z8-25.8和Z8-25.9之间110kb的区域内。该区间内存在1个与玉米甜质基因Sugary 1氨基酸序列相似性高达82.2%的基因LOC_Os08g40930,编码一个属于淀粉去分支酶（DBE）途径的异淀粉酶ISA1。测序结果表明,该基因序列和启动子在野生型和m5788中不存在碱基差异。qRT-PCR分析结果表明,与野生型相比,突变体中LOC_Os08g40930的表达量明显降低。同时,DBE途径中支链淀粉酶的编码基因表达量也显著降低。因此,m5788携带的isa1基因是一个新发现的等位变异。     

玉米粒长性状主基因+多基因遗传分析

玉米粒长是培育优良玉米品种的重要选择性状。选取粒长性状表现差异显著的玉米自交系铁7922和E28,及其组建的6个世代群体P₁、F₁、P₂、B₁、B₂和F₂为材料,运用主-多基因混合模型遗传分析方法进行分析,研究玉米粒长的遗传规律。结果表明：粒长性状在F₁表现为超亲优势,符合两对加性-显性-上位性主基因+加性-显性-上位性多基因遗传的E-1-0模型,主基因遗传率为41.22%~80.58%,多基因遗传率为17.68%~24.95%,环境因素决定粒长表型变异的19.42%~41.10%,控制玉米粒长性状的主基因效应高于多基因效应,并且主基因的加性累积效应明显,该性状在育种中可以通过世代累积进行选择。     

玉米雄性不育突变体mi-ms-3的遗传分析及分子鉴定

Maize is one of the best crops in the utilization of heterosis. Male sterile lines are important germplasms for the hybrids production. A male sterile mutant named mi-ms-3 was obtained by screening in a mutator insertion library. The number of male anthers in tassel decreased and not exserted. There were few anthers with only two pollen sacs in the mutant tassels, and some of the anthers were degenerated to membranous and formed filaments at their ends. Although pollens in the anthers could be stained by I₂-KI, pollen shedding was abnormal and the number of pollen grains decreased. The number of silks in the ear of the mutant increased, and there was a sterile grain on both sides of the maturated kernel. Fertility of F₁ plants, which were obtained by hybridization between mi-ms-3 and maize inbred Mo17, was normal. Genetic analysis of F₂ population showed that the mutant phenotype was controlled by a recessive gene. The candidate gene was preliminarily mapped on the long arm of chromosome 3 by BSA and it was located between a SSR marker and an Indel marker with a distance of 1.5 cM. There are 21 candidate genes in this region. It was finally found that the insertion mutation of Mu transposon occurred at 30 bp upstream of the coding region of zm00001d042618 (zmm16) by transponson tagging and sequencing analysis. The results showed that mi-ms-3 was a new allele of sts1, which caused by a single base mutation in the coding region. RT-PCR analysis indicated that the expression of zmm16 in the mutant was decreased. The identification of the new allelic mutant of sts1 in this study would provide new materials for the study of flower development and hybrid seed production.     

玉米tasselseed突变体ts12的遗传分析与分子鉴定

性别决定与玉米雄穗和雌穗发育密切相关,性别决定基因功能研究对性别决定分子机制的解析具有重要意义。利用甲基磺酸乙酯(EMS)处理B73花粉,获得了一个玉米雄穗结实突变体tasselseed12(ts12)。用扫描电镜对ts12突变体雄穗的形态学观察,发现未成熟雄穗长13 mm时,小穗呈现出明显的雌性化特征。利用图位克隆的方法,把ts12定位于分子标记LM4和RM5之间,物理距离约为290 kb,该区间共有9个注释基因,其中包括已报道的性别决定基因Tasselseed2(Ts2)。通过克隆ts12突变体中Ts2基因编码序列,发现Ts2基因编码区第196个碱基鸟嘌呤被替换为腺嘌呤,导致该位点编码的甘氨酸被替换为精氨酸,由此推测该保守位点突变可能是tasselseed表型产生的原因。ts12和ts2等位性测验结果表明所有F1、F2代植株雄穗均可产生花丝,推测ts12是ts2基因一个新的等位突变体。以加外源茉莉酸(JA,1 mmol L^-1)处理ts12突变体,发现处理后的小穗大部分可恢复正常。Ts2基因表达分析揭示在正常植株未成熟雄穗中的表达量最高,其次是未成熟雌穗及叶片中;在ts12未成熟雄穗和雌穗中,该基因的表达量极显著降低。Ts2保守位点的突变及其引起的表达量的降低可能是tasselseed表型产生的原因。     

玉米籽粒突变体smk7的表型分析和基因定位

利用甲基磺酸乙酯(EMS)对玉米自交系B73进行诱变,获得一个可以稳定遗传的小籽粒突变体smk7(small kernel 7)。smk7成熟籽粒表现为体积变小,胚和胚乳发育缺陷,百粒重显著降低。突变籽粒发芽率仅为10%,且幼苗黄化不能生长成正常植株。成熟smk7胚乳中淀粉、蛋白、油分含量与野生型籽粒相比无显著差异,但突变体胚乳淀粉粒体积明显变小且形状不规则。smk7突变籽粒在授粉后12 d即可观察到明显的小籽粒和空瘪表型,石蜡切片显微观察显示突变籽粒的胚和胚乳发育迟缓,胚乳基部转移层细胞(BETL)相对于野生型细胞壁向内生长减少,发育受阻。用杂合植株(+/smk7)与多个自交系分别杂交,构建不同背景的F2分离群体,遗传分析结果表明该性状受单隐性核基因控制。利用靶向测序基因型分型(genotyping by target sequencing,GBTS)技术将基因初定位于2号染色体短臂,进一步精细定位发现该基因位于RM1433917和RM1535316两个标记之间约120 kb的物理范围内,共有8个蛋白编码基因。本研究为进一步克隆和解析SMK7基因调控玉米籽粒发育的分子机制奠定了基础。     

一个新的玉米Vp15基因等位突变体的遗传分析与分子鉴定

在玉米繁种过程中发现一个玉米穗发芽突变体(viviparous), 命名为vp-like4。经过连续多代自交发现该突变体性状能稳定遗传, 并且表现为隐性单基因控制。以vp-like4与自交系Mo17杂交构建F₂遗传定位群体, 利用BSR-Seq方法, 将目的基因定位于玉米第5染色体173.8~175.6 Mb之间。通过基因组序列信息分析发现, 在此定位区间内存在一个已报道的Vp15基因。Vp15基因编码钼喋呤合酶小亚基, 参与类胡萝卜素裂解为ABA的过程。利用2个独立的vp15突变体vp15-umu1和vp15-DR1126的杂合体, 分别与vp-like4突变体杂合体做杂交进行等位测验, 发现杂交后代中正常籽粒与穗发芽籽粒比例符合3∶1分离比。基因组序列分析发现vp-like4突变体中Vp15基因在第2外显子末端及3°非翻译区有60个碱基的缺失, 与所报道的vp15突变体vp15-umu1、vp15-DR1126均在第2个外显子有Mutator转座子插入的突变方式不同。进一步通过RT-PCR检测发现, vp-like4突变体中Vp15基因的表达量显著降低。以上实验证据表明, vp-like4是一个新的Vp15基因等位突变体。