花生种子全长cDNA文库的构建和鉴定

以花生品种闽花5号各发育时期的种子为材料分离mRNA,利用SMART技术合成双链cDNA。利用限制性内切酶SfiⅠ酶切。将经过分级分离得到的cDNA连接到质粒载体pDNR-LIB,成功构建花生种子全长cDNA文库。将所得到初级文库扩增后进行保存,检测扩增文库滴度为1.7×109cfu/mL。PCR鉴定重组子,发现重组率接近100%,插入片段集中分布在0.5~2.0 kb。插入片段的平均大小在1000 bp左右,这说明该文库既可以满足低丰度基因的筛选,也可保证获得全长cDNA。     

巴西橡胶树低温诱导全长cDNA文库构建

通过低温诱导处理巴西橡胶树(Hevea brasiliensis)品系93-114幼嫩枝条和树叶,提取总RNA,并分离纯化mRNA.利用SMART技术合成橡胶树全长cDNA,经过Sfi I酶切后连接到pDNR-LIB载体中,得到全长cDNA文库.文库库容为1.4×106,重组率达到100%.挑取单菌落提取质粒并酶切鉴定,插入片断分布在0.3～2 kb之间,平均大小为1 kb左右,表明该文库质量合格,可用于全长基因的筛选.     

东乡野生稻育性恢复性的鉴定与遗传分析

以东乡野生稻水桃树、东塘上2个居群与不同细胞质来源的5个水稻不育系(B06S、珍汕97A、协青早A、中97A和粤泰A）配组,根据F1结实率的高低对东乡野生稻育性恢复性进行鉴定。建立了10个组合的P1、F1、P2、F2群体,利用混合模型理论的Akaike信息准则(AIC),在F2代中鉴定影响数量性状的主基因存在与否,主基因存在时通过分离分析估计主基因和微效基因的遗传效应以及所占总变异的分量。F1代的结实率变化范围为45.98%~76.57%,表明东乡野生稻具有一定的育性恢复性。F2代中该性状符合1对主基因+多基因的遗传模式,主基因遗传率为56.63%~88.29%,多基因遗传率为2.74%~30.97%,总基因型遗传率为63.17%~94.01%。中9A /东塘上居群杂交组合的F2代中,主基因是加性遗传,无显性效应,其他9个组合主基因是完全显性遗传。     

东乡野生稻研究进展及育种利用策略

东乡野生稻是目前世界上已发现的分布最北（N 28°14′、E 116°36′）的野生稻特异种质资源,其基因组中蕴含着抗病、抗虫、耐逆及其他诸多优良基因/QTL,前人已对其开展了大量的研究。本文从抗病、抗虫、抗逆、育性、产量及其他优良特性等方面总结了东乡野生稻的研究进展,并提出了其育种利用策略。     

东乡野生稻南昌异位圃内的开花习性

于2009年采用常规野外考察的方法,对南昌异位圃东乡野生稻的开花习性进行了调查。结果表明,南昌异位圃东乡野生稻开花时间比预期提前了近一个月,异位圃内各方向开花时间差异较大,异位圃南部开花高峰期在9月2日～9月4日,同一穗头上的可育度从上到下一般是逐渐递增,上部与下部育性差异达极显著,结实率40%左右。     

应用SMART技术构建甘蔗叶片全长cDNA文库

以甘蔗（Saccharum officinarum Linn）为材料,利用SMART技术构建甘蔗叶片全长cDNA文库,从甘蔗叶片中提取总RNA并分离mRNA,用分离到的微量mRNA合成cDNA第1链,通过长距离PCR扩增得到足量的双链cDNA,同时比较了不同双链cDNA与载体的连接效率和转化效率。结果表明,所构建的甘蔗叶片cDNA文库库容和重组百分比分别是1．8×106个克隆和98．6％,插入片段长度主要集中在0．6-2．5kb,平均插入片段长度约为1．2kb。本研究为国内首次报道甘蔗全长cDNA文库,所构建的文库拥有较好的质量,为项目组进一步开展甘蔗功能基因组学研究提供了平台。     

东乡野生稻的粒形相关QTL分析

以东乡野生稻/协青早B∥协青早B的回交自交系群体为遗传材料,利用简化基因组测序技术鉴定群体的SNP分子标记基因型,对东乡野生稻的粒形相关性状进行QTL分析。在水稻第1,3,4,5,10,12号染色体上共发现14个粒形相关QTL,包括6个粒长QTL(q GL1、q GL3.1、q GL3.2、q GL4、q GL10.1、q GL10.2)、2个粒厚QTL(q GT5、q GT10)和6个粒重QTL(q KGW3、q KGM4、q KGW5、q KGW 10.1、q KGW10.2、q KGW12)。其中q GL3.1与q KGW3、q GT5与q KGW5、q GL4与q KGM4、q GL10.1与q KGW10.1以及q GL10.2、q GT10与q KGW10.2定位区间重叠,推测这些QTL具有一因多效特性。加性效应最大的粒长QTL即q GL3.1(q KGW3)被定位在第3染色体的粒形基因GS3的区域中,且q GL3.1与GS3都是控制粒长的主效QTL,推测q GL3.1就是GS3基因。而在q GL3.1区域附近还存在粒长QTL q GL3.2,该位点与已克隆的粒长QTL即GL3.1在位置上重叠,推测q GL3.2可能就是GL3.1/Os PPKL1基因。     

东乡野生稻有利性状鉴定与育种利用研究进展

东乡野生稻具有丰富的遗传多样性,是水稻育种的宝贵资源。综述了东乡野生稻高产性状、抗病性、抗虫性、耐寒性、抗旱性、细胞质雄性不育及其育性恢复和其它优良性状的鉴定、基因定位、遗传及其育种利用等方面的研究进展,并就进一步研究提出了展望。     

东乡野生稻种质资源保护现状、存在问题及对策

通过对江西省东乡野生稻种质资源的野外调查,结合东乡野生稻原生境保护区实地监测结果,分析了东乡野生稻种质资源保护现状及其存在的主要问题,提出了进一步保护和利用的对策,为更好保护东乡野生稻种质资源和研究利用提供借鉴。     

Identification and Characterization of Drought Stress- Responsive Novel microRNAs in Dongxiang Wild Rice

MicroRNAs(miRNAs) are non-coding small RNAs, which play important regulatory roles in response to biotic and abiotic stresses. Dongxiang wild rice(Oryza rufipogon, DXWR) can survive in extreme drought environment, but its molecular mechanism of drought resistance is still largely unknown. To further explore miRNA regulatory mechanisms involved in drought resistance, we identified 138 novel miRNAs in DXWR using small RNA sequencing and bioinformatics approaches, and found that the expression levels of 67 novel miRNAs were significantly affected by drought stress. In total, 200 candidate target genes were predicted and annotated for the drought stress-responsive novel miRNAs. Gene Ontology(GO) analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathways suggested that most of the target genes were related to metabolism. Stem-loop quantitative real-time PCR(qRT-PCR) results exhibited high concordance with sequencing data, which confirmed that miRNA expression patterns based on small RNA sequencing in the present study were reliable. Meanwhile, qRT-PCR validated the inverse expression patterns between several miRNAs and their target genes. These results will enhance our understanding of miRNA regulatory mechanisms in response to drought stress in DXWR, and can serve as an important reference for the protection and utilization of this valuable genetic resource.     

Identification and Genetic Analysis of Fertility Restoration Ability in Dongxiang Wild Rice(Oryza rufipogon)

Two populations of Dongxiang wild rice(Oryza rufipogon),Shuitaoshu and Dongtangshang,were crossed with five male sterile rice lines with different cytoplasmic backgrounds(B06S,Zhenshan 97A,Xieqingzao A,Zhong 9A,and Yuetai A),and the seed setting rate of the F1 was used to judge the fertility restoration ability in the Dongxiang wild rice.With P1.F1,P2,and F2 populations as materials,the Akaike's Information Criterion(AIC)was used to identify the major genes affecting quantitative traits,and when the major genes existed,the genetic effects of the major gene and the polygene and their genetic variance were estimated through segregation analysis.The seed setting rates of the F1 generation varied from 45.98%to 76.57%,suggesting that the Dongxiang wild rice had the fertility restoration ability.One major gone plus polygene mixed inheritance model was the most fitted genetic model for this trait in all the F2 populations.The heritability values of the major genes varied from 56.63%to 88.29%and those of the polygenes from 2.74%to 30.97%,and the total heritability values were from 63.1 7%to 94.01%.The major gone inheritance of the combination Zhong 9A/Dongtangshang was controlled by the additive effect without dominant effect,and the other nine combinations were by the completely dominant inheritance.     

Identification and Genetic Analysis of Fertility Restoration Ability in Dongxiang Wild Rice （Oryza rufipogon）

Two populations of Dongxiang wild rice (Oryza rufipogon), Shuitaoshu and Dongtangshang, were crossed with five male sterile rice lines with different cytoplasmic backgrounds (B06S, Zhenshan 97A, Xieqingzao A, Zhong 9A, and Yuetai A), and the seed setting rate of the F₁ was used to judge the fertility restoration ability in the Dongxiang wild rice. With P₁, F₁, P₂, and F₂ populations as materials, the Akaike's Information Criterion (AIC) was used to identify the major genes affecting quantitative traits, and when the major genes existed, the genetic effects of the major gene and the polygene and their genetic variance were estimated through segregation analysis. The seed setting rates of the F₁ generation varied from 45.98% to 76.57%, suggesting that the Dongxiang wild rice had the fertility restoration ability. One major gene plus polygene mixed inheritance model was the most fitted genetic model for this trait in all the F₂ populations. The heritability values of the major genes varied from 56.63% to 88.29% and those of the polygenes from 2.74% to 30.97%, and the total heritability values were from 63.17% to 94.01%. The major gene inheritance of the combination Zhong 9A/Dongtangshang was controlled by the additive effect without dominant effect, and the other nine combinations were by the completely dominant inheritance.     

东乡野生稻重要耐逆性状及其分子机制研究进展

东乡野生稻是全球分布最北的普通野生稻,具有丰富的抗逆性状,其育种利用价值高。本文综述了近年来东乡野生稻在耐冷、耐旱及抗虫等重要耐逆性状及其分子机制研究方面取得的进展,以期为水稻耐逆性的进一步研究提供依据。