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Chinese Science Bulletin 2007年第17期52卷 2358-2364页

作者： PAN YuXin MA Jun ZHANG GuiYin HAN GaiYing WANG XingFent MA ZhiYingt Key Laboratory of Crop Germplasm Resources of Hebei Province Agricultural University of Hebei Baoding 071001 China

Cotton fiber strength is mainly determined during the secondary cell wall deposition stage when cel- lulose is synthesized. We obtained cdna of 20―25 d post anthesis (DPA) fiber from 109 F2 progeny and developed a cotton fiber transcriptome profiling via cdna-aflp technology using 37 different primer combinations. The F2 population originated from an interspecific cross between Gossypium hirsutum and Gossypium barbadense. One hundred and thirty-eight absence/presence polymorphic transcript- derived fragments (TDFs), with sizes ranging from 100 bp to 722 bp, were screened. Of these, 75 (53.62%) were polymorphic between the parents of the F2 population. Sequencing the 75 transcripts revealed that 37 of them had been reported to be cotton fiber ESTs. Nine of 75 transcript sequences were homologous to 7 cloned cotton fiber genes, encoding cysteine proteinase, vacuolar H+-pyro- phosphatase, vacuolar H+-ATPase, catalytic subunit, arabinogalactan protein, putative receptor protein kinase PERK1, GIA/RGA-like gibberellin response modulator and cellulose synthase. Some other transcripts may represent new gene fragments in cotton fiber development. Surprisingly, 46 of the 75 transcripts were mapped to a single linkage group. The transcriptome groups and the sequenced TDFs could serve as important resources in the functional genomic research of cotton fiber development.

关键词：棉花 cdna-aflp 基因转录纤维生长细胞壁

cdna-aflp analysis reveals that maize resistance to Bipolaris maydis is associated with the induction of multiple defense-related genes

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Chinese Science Bulletin 2001年第17期46卷 1454-1458页

作者： GAO Zhihuan XUE Yongbiao DAI Jingrui Department of Plant Genetics and Breeding China Agricultural UniversityBeijing 100094China Laboratory of Plant Genetics and Developmental Biology Institute of Developmental BiologyChinese Academy of SciencesBeijing 100080China

The fungal pathogen Bipolaris maydis invades by direct penetration into maize leaf veins. In order to understand the resistance mechanism of maize to B. maydis strain 523, cdna-aflp (amplified fragment length polymorphism) analysis was conducted to compare the changes in mRNA transcripts hi response to B. maydis infection between a highly disease-resistant (HDR) line and a susceptible (S) line. 13 cdna fragments derived from the genes showing enhanced expression after fungal infection, named HDR genes, were isolated from the HDR line. Northern blot analysis showed that 5 HDR genes were induced by fungal infection in the HDR, but not the S lines. The 5 HDR genes showed homology to previously characterized genes involved in disease resistance. A full-length HDR10 cdna was isolated. It had a capacity to encode a protein of 284 amino acids. The deduced amino acid sequence of the HDR10 gene was homologous to a fungal infection-induced protein from Cicer arietinum and a hypersensitive response protein from

关键词： maize Bipolaris maydis cdna-aflp plant defense gene expression.

Diversity and Genetic Differentiation of the Whitefly Bemisia tabaci Species Complex in China Based on mtCOI and cdna-aflp Analysis

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Journal of Integrative Agriculture 2012年第2期11卷 206-214页

作者： GUO Xiao-jun RAO Qiong ZHANG Fan LUO Chen ZHANG Hong-yu GAO Xi-wu Department of Entomology China Agricultural University Beijing 100193 P.R.China Institute of Plant and Environmental Protection Beijing Academy of Agriculture and Forestry Sciences Beijing 100097 P.R. China College of Plant Science and Technology Huazhong Agricultural University Wuhan 430070 PR. China

The whitefly Bemisia tabaci are considered as a taxonomically complex that contained some destructive *** of the most prevalent cryptic species are *** Middle East-Asia Minor 1（MEAM1）and Mediterranean（MED）.In an extensive field survey of the *** complex present throughout part of China from 2004 to 2007,we obtained 93 samples of *** from 22 *** determined that these Chinese haplotypes included 2 invasive species（MEAM1 and MED）,and 4 indigenous cryptic species（Asia II 1,Asia II 3,China 3 and Asia II 7）by sequencing mitochondrial cytochrome oxidose one gene（mtCOI）.The diversity and genetic differentiation of a subset of 19 populations of *** were studied using cdna amplified fragment length polymorphism（aflp）.Prior to 2007,MEAM1 was a dominant species in many provinces in *** 2007,MED was dominant in 11 *** invasive and indigenous species were simultaneously found in some *** species of *** were found in six provinces in southern *** and MEAM1 have broad ranges of host plants,and indigenous species appeared to have much narrower host *** Asia II 3 samples were found on cotton except one on *** 3 has more host plants than Asia II *** samples of China 3 were collected from sweet potato,Japanese hop,squash and cotton.A total of 677 reproducible bands amplified with 5 aflp primer combinations were *** highest proportion of polymorphic bands was 98.7% and the lowest was 91.9%.Unweighted pair-group method analysis indicated that the clustering was independent of the different *** showed the lowest degree of similarity than the other *** data indicate that both MEAM1and MED were rapidly established in China.

关键词： Bemisia tabaci mtCOI cdna-aflp diversity whitefly

Relationship between differ- ential gene expression pat- terns in functional leaves of maize (Zea mays L.) at milk filling stage and heterosis using cdna-aflp

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Chinese Science Bulletin 2003年第1期48卷 76-81页

作者： TIAN Zengyuan & DAI Jingrui National Maize Improvement Center of China, China Agricultural Uni-versity, Beijing 100094, China Correspondence should be addressed to Tian Zengyuan (e-mail: tian-zengyuan@***) National Maize Improvement Center of China China Agricultural University Beijing China

To understand the molecular mechanism of maize heterosis, differential gene expression patterns in the functional leaves of 35 maize hybrids relative to their parents involving 10 elite inbreds at milk filling stage were analyzed by using cdna-aflp. The correlation analyses of various differential expression patterns with the performance and heterosis of main maize agronomic traits were evaluated. The main results were as follows: For uniparental specific expression, significant positive correlations were detected with the performance of seed weight per ear and 100-seed weight at 0.01 and 0.05 probability levels respectively. For biparental specific expression, significant negative correla-tions were detected with the performance of ear diameter and seed weight per ear at 0.01 probability level. For uni-parental specific expression, significant positive correlations were detected with the heterosis of ear diameter and seed weight per ear at 0.01 and 0.05 probability levels respectively. For biparental specific expression, significant negative cor-relation was detected with the heterosis of ear diameter at 0.05 probability level. However, for F1-specific expression, for fragments detected only in one parent and F1, and for fragments detected only in two parents or only in F1, no sig-nificant correlation was detected with the performance or heterosis of all agronomic traits surveyed.

关键词：玉米灌浆期叶片差异基因表达模式杂种优势相关性 cdna-aflp 农艺性状

Identification of genes induced during Medicago sativa nodule development by using the cdna-aflp technique

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Chinese Science Bulletin 2006年第17期51卷 2087-2094页

作者： XIE Rong HE Zhishui LIU Fanghua ZOU Huasong ZHU Jiabi YU Guanqiao National Laboratory of Plant Molecular Genetics Institute of Plant Physiology & Ecology Shanghai Institutes for Biological SciencesChinese Academy of Sciences Shanghai 200032 China Graduate School of the Chinese Academy of Sciences Beijing 100039China

A variety of genes involved in nodule development and function were induced during sym biotic root nodule development in alfalfa. We identi fied those genes by comparing the cdna-amplified fragment length polymorphism (cdna-aflp) pat terns of infected nodules and uninfected roots. Sev enty-one transcript-derived fragments (TDFs) were isolated from about 3000 TDFs, which are enhanced or specifically expressed in nodule. The differentia expression patterns of 61 genes were confirmed b reverse Northern hybridization or Northern hybridiza tion analyses. Among these, 31 exhibited significan similarities to characterized database entries. These results suggest that the genes corresponding to the 31 TDFs may play important roles in signal transduc tion, gene expression and regulation, substance transportation or auto-regulation of nodulation during nodule development. The Northern hybridization analysis also indicates that the gene corresponding to TDF RX89 was not expressed in the nodule elic ited by bacA mutant, presumably it is specially in duced at the stage of nodule development when bacteroids begin to be differentiated.

关键词：多态性 cdna 节结共生固氮作用 cdna-aflp

Chinese Cabbage-pak-choi Transcriptome Map Construction with cdna-aflp Techniques

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Agricultural Sciences in China 2008年第10期7卷 1181-1188页

作者： FAN Shu-ying LE Jian-gang CHENG Guang-jie WU Cai-jun College of Agronomy Jiangxi Agricultural University Nanchang 330045 P.R.China

Chinese cabbage-pak-choi （Brassica campestris L. ssp. chinensis Makino） transcriptome map with cdna-aflp techniques was constructed. The inbred line Aijiaohuang 97-3-2, the inbred line Baimanjing 001-24 of turnip [B. campestris ssp. rapifera （Matzg.） Sinsk] and 183 F6 （recombinant inbred population） plants were used as experimental materials, cdnas were synthesized from total RNA extracted from young leaves at rosette stage. 256 pairs of cdna-aflp primers were used to detect the polymorphisms between parents Aijiaohuang 97-3-2 and Baimanjing 001-24. 56 pairs of cdna-aflp primers with high polymorphisms were screened from 256 pairs of primer by DNA-aflp techniques. The genetic diversity of parents and 183 F6 progenies was detected by 56 pairs of cdna-aflp primers. The segregation and distribution of cdna-aflps molecular marker were analyzed to construct transcriptome map amongst parents and F6 plants. A total of 164 cdna-aflps marker loci were mapped into 13 linkage groups which covered 1401.2 cM with an average distance of 9.7 cM. It was the first transcriptome map of Chinese cabbage using cdna-aflp technique.

关键词： Brassica campestris ssp. chinensis cdna-aflp transcriptome map

Construction of a cdna-aflp reaction system of Populus euphratica for abiotic stress studies

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Forestry Studies in China 2011年第2期13卷 104-111页

作者： MA Hong-shuang XIA Xin-li YIN Wei-lun National Engineering Laboratory for Tree Breeding Beijing Forestry University Beijing 100083 E R. China Preparation Office of Taiyuan Taishan Botanical Garden Taiyuan 030006 P. R. China

We established a cdna-aflp （amplified fragment length polymorphism） system for Populus euphratica, after optimizing several key factors. The results show that optimized CTAB method is suitable for the extraction of ideal RNA from P euphratica; the double-stranded cdna （ds-cdna） is completely digested by the combination of EcoRI and MseI for 4.5 h; the reducible result can be obtained when the preamplification products were diluted 10-fold for selective amplification templates. The amplification products resulting from this reaction system were determined by silver staining and the fragments were clear and stable. Detected by RT-PCR, the optimized differential display cdna-aflp technique can accurately reflect the true differences of gene expressions among plants. In addition, 42 pairs of primer arrays, producing repeatable and prolific bands, were successfully selected by this improved method. This investigation establishes a cdna-aflp system for P. euphratica, which forms a basis for further study on stress-response genes in P. euphratica.

关键词： Populus euphratica cdna-aflp abiotic stress mechanism

利用cdna-aflp技术分析炭疽菌危害诱导茶树的差异表达基因

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茶叶科学 2020年第1期40卷 26-38页

作者：魏日凤赖建东彭成彬张承康连玲丽刘伟福建农林大学园艺学院福建福州350002 宁德师范学院生命科学学院福建宁德352100 福建农林大学生命科学学院福建福州350002 湖北科技学院湖北咸宁437100

应用cdna-aflp技术筛选炭疽菌(Colletotrichum sp.1)危害诱导茶树毛蟹品种(Camellia sinensis cv. Maoxie)的差异表达基因,为探究茶树抗炭疽菌的分子机制奠定基础。利用256对选择性扩增引物组合分析病菌接种后0 h和48 h的叶片cdna,经筛... 详细信息

应用cdna-aflp技术筛选炭疽菌(Colletotrichum sp.1)危害诱导茶树毛蟹品种(Camellia sinensis cv. Maoxie)的差异表达基因,为探究茶树抗炭疽菌的分子机制奠定基础。利用256对选择性扩增引物组合分析病菌接种后0 h和48 h的叶片cdna,经筛选、测序和BLAST比对获得136条差异表达片段(TDFs)。同源基因功能分析结果表明,128条TDFs与Nr数据库已有基因同源,其功能以胁迫响应、生物调控与信号转导为主;其中51条TDFs与TPIA数据库中冷害、干旱、盐碱条件下的差异表达基因高度同源。进一步对27条TDFs进行qRT-PCR验证,结果显示,有24条TDFs的表达情况与cdna-aflp的结果一致,WRKY转录因子、乙烯响应转录因子、过氧化物酶和超氧化物歧化酶等抗逆相关基因的表达显著上调。本试验通过筛选获得差异表达的抗逆相关基因,为后续的基因功能研究奠定基础。

关键词：茶树炭疽菌 cdna-aflp 差异表达基因

TMV侵染烟草基因差异表达的cdna-aflp分析

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作物学报 2012年第1期38卷 62-70页

作者：陈荣平刘磊万秀清邱恩建王春军宋宝刚颜培强杨铁钊河南农业大学烟草学院河南郑州450002 黑龙江省烟草公司牡丹江烟草科学研究所黑龙江牡丹江157011 牡丹江市疾病防控中心黑龙江牡丹江157022

以烤烟品种龙江925[高抗烟草普通花叶病(TMV)]接种TMV的叶片为材料,选用240对引物组合的扩增,获得约9500个转录基因片段,经过克隆测序最终获得了12个诱导表达片段,它们与核酸代谢、蛋白质合成与修饰、能量代谢、胁迫响应、细胞内运输、... 详细信息

以烤烟品种龙江925[高抗烟草普通花叶病(TMV)]接种TMV的叶片为材料,选用240对引物组合的扩增,获得约9500个转录基因片段,经过克隆测序最终获得了12个诱导表达片段,它们与核酸代谢、蛋白质合成与修饰、能量代谢、胁迫响应、细胞内运输、糖代谢等相关。利用荧光定量PCR分析诱导表达片段TIF2在0～72h之间5个时间点(0、12、24、48和72h)的基因差异表达,证实了所获得的基因序列为真实诱导的表达序列。对此序列进行RACE,最终获得全长cdna序列,全序列857bp,预测的编码区在101～613bp之间,共编码170个氨基酸。经Blastn、Blastp比对分析,在烟草中没有获得其同源序列,确定该基因是在烟草中发现的与抗TMV相关的新基因。

关键词：烟草 TMV 差异基因表达 cdna-aflp

利用cdna-aflp技术分析‘窄冠黑白杨’杂种优势的差异表达基因

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植物生理学报 2020年第12期56卷 2695-2704页

作者：侯丽丽姚培娟李际红李景涛李柱山东农业大学林学院/泰山森林生态系统国家定位观测研究站/黄河下游森林培育国家林业和草原局重点实验室山东泰安271018 山东省林木种苗和花卉站济南250014 山东农业大学科研处山东泰安271018

本研究以‘窄冠黑白杨’(Populus ‘Zhaiguanheibaiyang’)及其亲本为材料,通过cdna扩增片段长度多态性分析技术(cdna-aflp)构建扩增图谱。从96对扩增引物中筛选出35对可用于条带分析的扩增引物,获得761个修饰位点, 5种基因差异表达模... 详细信息

本研究以‘窄冠黑白杨’(Populus ‘Zhaiguanheibaiyang’)及其亲本为材料,通过cdna扩增片段长度多态性分析技术(cdna-aflp)构建扩增图谱。从96对扩增引物中筛选出35对可用于条带分析的扩增引物,获得761个修饰位点, 5种基因差异表达模式。回收cdna-aflp图谱中分离的19条差异表达的转录衍生片段(transcript-derived fragments, TDFs)进行同源性比对分析,发现其中14条TDFs与杨树基因库中的已知基因同源,如丝氨酸/苏氨酸蛋白激酶(STPK)、泛素激活酶(E1)、糖基转移酶(UDP)、ARGOS-like蛋白基因等,在植物生长代谢活动、信号转导调控等途径中具有重要作用。利用实时荧光定量PCR技术检测差异表达TDFs在杂种及父本中芽、叶、茎等不同组织的表达量,发现在叶中表达量有明显差异,这可能与杂种优势密切相关。结果与cdna-aflp图谱一致,说明研究结果稳定可靠,为后续杨树杂种优势的研究提供新的见解。

关键词： ‘窄冠黑白杨’ 杂种优势 cdna-aflp 差异基因