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Construction and genetic analysis of mutator insertion mutant population in maize

Chinese Science Bulletin 2006年第21期51卷 2604-2610页

作者： LIU Wenting GAO Youjun TENG Feng SHI Qing ZHENG Yonglian National key laboratory of crop genetic improvement Huazhong Agricultural University Wuhan 430070 China

A total of 26718 M1 plants were ob- tained by crossing the active mutator transposon donor parents (Q105, WW51, 115F, V26-2 and 919J) with the recipient parents (Hz85,W328 with Bz gene and S-Mo17Rf3Rf3). The phenotypes of M1 plants were observed in the field. M1 plants were self-pollinated to develop the mutator insertion-mutagenized M2 seeds. The transposition frequency of the mutator in the genome was calculated based on the spotted aleurone phenotype of the M2 seeds. The results showed that: (1) the mutation frequency of M1 phe- notypes in the field was 0.07 in the population of W328×Mu; (2) the mutation frequency of spotted aleurone seeds on the M2 ears was 0.122 in the population of W328×Mu; (3) five S-cytoplasm male-sterile plants were found among 22500 M1 plants of S-Mo17Rf3Rf3×Mu, with the transposition frequency about 2.2×10?4 per locus. 99 flanking se- quences of mutator transposition were amplified by the modified MuTAIL-PCR, and 59 non-redundant sequences with length around 400 bp were obtained. After bioinformatic analysis, 27 sequences of them could be annotated, using non-redundant nucleotide database of maize, rice, and Arabidopsis. 36 se- quences of them were located on the genetic map of maize by comparative genomics, and several flank- ing sequences of mutator insertion were mapped on the single marker locus. Hotspot sequences of mu- tator transposition were revealed by comparing the homologies between the 9-bp target site duplication of the mutator insertion. The putative functions of 8 flanking sequences of mutator transposition had identity with the functions of their correspondingmarker. The constructed mutator insertion mutant population in maize will facilitate the new gene discovery and functional genomics study in maize.

关键词：玉米突变基因转位子突变体种群 MuTAIL-PCR

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Seed-specific overexpression of cotton GhDGAT1 gene leads to increased oil accumulation in cottonseed

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The crop Journal 2021年第2期9卷 487-490页

作者： Peng Wu Xiaolan Xu Jingwen Li Jun Zhang Siyuan Chang Xiyan Yang Xiaoping Guo National key laboratory of crop genetic improvement Huazhong Agricultural UniversityWuhan 430070HubeiChina

As the main byproduct of cotton fiber,the cotton seed yields about 1.6 times that of fiber,with its oil rich in unsaturated fatty acids,mainly linoleic *** is desirable for breeders to increase the oil content of cottonseed without affecting the yield and quality of cotton *** this study,a seed-specific promoter-(alpha-globulin gene promoter-)driven GhDGAT1 overexpression vector(P_(aGlob)-GhDGAT1)was constructed and used to transform an upland cotton line YZ1(Gossypium hirsutum).Overexpression of the cotton gene GhDGAT1 in cotton seeds increased its total oil content from 4.7%to 13.9%in different transgenic lines and different *** the increase of oil content,the composition and contents of the main fatty acids in cotton seed also changed,as reflected by the contents of the main saturated fatty acids and unsaturated oleic ***1 could be used to increase oil content and improve oil composition in cottonseed.

关键词： aGP promoter Cotton Fatty acid GhDGAT1 Seed oil

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Analysis of genes differentially expressed during initial cellular dedifferentiation in cotton

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Chinese Science Bulletin 2008年第23期53卷 3666-3676页

作者： ZHU HuaGuo TU LiLi JIN ShuangXia XU Li TAN JiaFu DENG FengLin ZHANG XianLong National key laboratory of crop genetic improvement Huazhong Agricultural University Wuhan 430070 China

The early phase of phytohormone induction is a vital stage of somatic embryogenesis. This phase includes a key process for acquiring cellular totipotency through cellular dedifferentiation. To unravel the molecular mechanism of cellular dedifferentiation in cotton, we constructed a cDNA library using the suppression subtractive hybridization method. A total of 286 differential cDNA clones were sequenced and identified. Among these clones, 112 unique ESTs were significantly up-regulated during the early phase of phytohormone induction, and 40.2% of the ESTs were first identified. GST was highly expressed from 6 to 24 h after induction with phytohormone treatment. PRPs were predominantly expressed and exhibited distinct expression patterns in different treatments, suggesting that they are closely related to cellular dedifferentiation in cotton. Putative GhSAMS, GhSAMDC, GhSAHH and GhACO3 involvement in SAM metabolism was identified in this library. The analysis of qRT-PCR showed that two remarkable increased expressions of the four SAM-related genes happened during the early phase of phytohormone induction, and that a highly positive correlation existed between GhSAMS and GhSAHH. The highest expression level of GhSAMS might be associated with its reentry into the cell cycle. The histological observations further showed that some cells accomplished cellular dedifferentiation and division within 72 h in 2,4-D treatment, and that cellular dedifferentiation might be regulated through two alterations in SAM-dependent transmethylation activity in cotton. In addition, the expression patterns of differential genes in different treatments disclosed the complicated interaction between 2, 4-D and kinetin.

关键词：细胞去分化棉花脯氨酸蛋白质转甲基作用

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Studies of new EST-SSRs derived from Gossypium barbadense

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Chinese Science Bulletin 2007年第18期52卷 2522-2531页

作者： ZHANG YanXin LIN ZhongXu LI Wu TU LiLi NIE YiChun ZHANG XianLong National key laboratory of crop genetic improvement Huazhong Agricultural University Wuhan 430070 China

Existing cotton EST-SSR markers are mostly derived from Gossypium arboreum and Gossypium hir-sutum, but EST-SSR markers from Gossypium barbadense are scarce. One hundred and nineteen EST-SSRs were developed based on 98 unique ESTs from a cDNA library constructed in our laboratory using developing fibers from G. barbadense cv. Pima3-79. Among the SSRs, trinucleotide AAG appeared at a high frequency of 11.76%. 36 accessions (consisting of 13 diploids of the A genome, 11 diploids of the D genome and 12 allotetraploids of the AD genome) were employed to test new EST-SSRs. 76 EST-SSRs were successfully amplified, and 313 polymorphic fragments were yielded, with an average of 4.11 fragments per primer pair. The PIC ranged from 0.17 to 0.95 with an average of 0.53. Based on Jaccard’s genetic similarity coefficient, these 36 accessions were clustered into three groups. 21 EST-SSRs exhibited polymorphisms in BC1 population ((Emian22 × Pima3-79) × Emian22), 24 polymor- phic loci were generated, while 22 of the 24 polymorphic loci were integrated with our interspecific BC1 backbone genetic linkage map, and anchored in 12 chromosomes. This study effectively proved that EST-SSRs from G. barbadense are valuable for genetic diversity analysis and genetic mapping.

关键词：棉花基因表达基因序列分析遗传多样性基因图谱

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OcBSA:An NGS-based bulk segregant analysis tool for outcross populations

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Molecular Plant 2024年第4期17卷 648-657页

作者： Lingkui Zhang Yanfeng Duan Zewei Zhang Lei Zhang Shumin Chen Chengcheng Cai Shaoguang Duan Kang Zhang Guangcun Li Feng Cheng State key laboratory of Vegetable Biobreeding Key Laboratory of Biology and Genetic Improvement of Horticultural Crops of the Ministry of Agriculture and Rural AffairsKey Laboratory of Biology and Genetic Improvement of Tuber and Root Crop of Ministry of Agriculture and Rural AffairsSino-Dutch Joint Laboratory of Horticultural GenomicsInstitute of Vegetables and FlowersChinese Academy of Agricultural SciencesBeijing 100081China

Constructing inbred lines for self-incompatible species and species with long generation times is challenging,making the use of F1 outcross/segregating populations the main strategy for genetic studies of such ***,there is a lack of dedicated algorithms/tools for rapid quantitative trait locus(QTL)mapping using the F1 *** this end,we have designed and developed an algorithm/tool called OcBSA specifically for QTL mapping of F1 *** transforms the four-haplotype inheritance problem from the two heterozygous diploid parents of the F1 population into the two-haplotype inheritance problem common in current genetic studies by removing the two haplotypes from the heterozygous parent that do not contribute to phenotype segregation in the F1 *** of OcBSA on 1800 simulated F1 populations demonstrated its advantages over other currently available tools in terms of sensitivity and *** addition,the broad applicability of OcBSA was validated by QTL mapping using seven reported F1 populations of apple,pear,peach,citrus,grape,tea,and *** also used OcBSA to map the QTL for flower color in a newly constructed F1 population of potato generated in this *** OcBSA mapping result was verified by the insertion or deletion markers to be consistent with a previously reported locus harboring the ANTHOCYANIN 2 gene,which regulates potato flower *** together,these results highlight the power and broad utility of OcBSA for QTL mapping using F1 populations and thus a great potential for functional gene mining in outcrossing *** ease of use,we have developed both Windows and Linux versions of OcBSA,which are freely available at:https://***/Bioinformaticslab/OcBSA.

关键词： outcross BSA self-incompatible F1 population QTL mapping potato

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Improving crop drought resistance with plant growth regulators and rhizobacteria: Mechanisms, applications, and perspectives

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Plant Communications 2022年第1期3卷 21-35页

作者： Hui Zhang Xiaopeng Sun Mingqiu Dai National key laboratory of crop genetic improvement Huazhong Agricultural UniversityWuhan 430070China

Drought is one of the main abiotic stresses that cause crop yield *** crop yield under drought stress is a major goal of crop breeding,as it is critical to food *** mechanism of plant drought resistance has been well studied,and diverse drought resistance genes have been identified in recent years,but transferring this knowledge from the laboratory to field production remains a significant ***,some new strategies have become research frontiers owing to their advantages of low cost,convenience,strong field operability,and/or environmental *** plant growth regulator(PGR)treatment and microbe-based plant biotechnology have been used to effectively improve crop drought tolerance and preserve yield under drought ***,our understanding of the mechanisms by which PGRs regulate plant drought resistance and of plant-microbiome interactions under drought is still *** this review,we summarize these two strategies reported in recent studies,focusing on the mechanisms by which these exogenous treatments regulate crop drought ***,future challenges and directions in crop drought resistance breeding are discussed.

关键词： abiotic stress drought tolerance exogenous plant growth regulators microbial crops

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An in planta haploid induction system in Brassica napus

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Journal of Integrative Plant Biology 2022年第6期64卷 1140-1144页

作者： Yifan Li Dan Li Qing Xiao Huadong Wang Jing Wen Jinxing Tu Jinxiong Shen Tingdong Fu Bin Yi National key laboratory of crop genetic improvement National Center of Rapeseed ImprovementHuazhong Agricultural UniversityWuhan 430070China key laboratory of crop Physiology Ecology and Genetic BreedingMinistry of EducationJiangxi Agricultural UniversityNanchang 330045China

Doubled haploid technology is widely used to accelerate plant breeding,but its use in the important oilseed crop Brassica napus *** limited because *** haploids could only be obtained through in vitro anther or microspore ***,maize(Zea mays)lines containing mutations in Domain of unknown function 679 membrane protein(DMP)were used as haploid inducer *** new haploid induction mechanism has been extended to several other plants,including the dicots Arabidopsis thaliana,tomato(Solanum lycopersicum),and tobacco(Nicotiana tabacum).Here,we knocked out four BnaDMP genes in the *** cultivar Westar using a clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9 vector with an enhanced green fluorescent protein expression *** with DMP mutations in *** in the T_(0),T_(1),and T_(2) generations exhibited a haploid induction rate up to 2.53%.These results suggest that targeting BnaDMP could be useful for haploid induction in ***.

关键词： Brassica napus CRISPR/Cas9 fluorescence screening maternal haploid induction

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Suitable internal control genes for qRT-PCR normalization in cotton fiber development and somatic embryogenesis

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Chinese Science Bulletin 2007年第22期52卷 3110-3117页

作者： TU LiLi ZHANG XianLong LIU DiQiu JIN ShuangXia CAO JingLin ZHU LongFu DENG FengLin TAN JiaFu ZHANG CunBin National key laboratory of crop genetic improvement Huazhong Agricultural University Wuhan 430070 China

The mechanisms of cotton fiber development and somatic embryogenesis have been explored sys-tematically with microarray and suppression subtractive hybridization. Real-time RT-PCR provides the simultaneous measurement of gene expression in many different samples,with which the data from microarray or others can be confirmed in detail. To achieve accurate and reliable gene expression re-sults,normalization of real-time PCR data against one or several internal control genes is required,which should not fluctuate in different tissues during various stages of development. We assessed the gene expression of 7 frequently used housekeeping genes,including 18S rRNA,Histone3,UBQ7,Actin,Cyclophilin,Gbpolyubiquitin-1 and Gbpolyubiquitin-2,in a diverse set of 21 cotton samples. For fiber developmental series the expression of all housekeeping genes had the same down tendency after 17 DPA. But the expression of the AGP gene(arabinogalactan protein) that has high expression level at the later fiber development stage was up-regulated from 15 to 27 DPA. So the relative absolute quanti-fication should be an efficient and convenient method for the fiber developmental series. The expres-sion of nonfiber tissues series varied not so much against the fiber developmental series. And three best control genes Histone3,UBQ7 and Gbpolyubiquitin-1 have to be used in a combinated way to get better normalization.

关键词：棉花纤维胚胎测量模式

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AFLP Fingerprinting and genetic Diversity of Main Sweetpotato Varieties in China

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Journal of Integrative Agriculture 2012年第9期11卷 1424-1433页

作者： LIU De-gao ZHAO Ning ZHAI Hong YU Xiao-xia JIE Qin WANG Lian-jun HE Shao-zhen LIUQing-chang Beijing key laboratory of crop genetic improvement/laboratory of crop Heterosis and Utilization Ministry of Education/ChinaAgricultural University Beijing 100193 P.R.China

AFLP fingerprinting of the 98 main sweetpotato varieties planted in China has been constructed. Using 17 AFLP primer combinations which were selected from 1 208 primer combinations and generated the most amounts of polymorphic bands, AFLP analysis of the 98 main sweetpotato varieties gave a total of 410 clear polymorphic bands with an average of 24.12 polymorphic bands per primer combination. Each one of the 98 sweetpotato varieties could be clearly distinguished by EcoR I-cta/Mse I-ggc primer combination which generated the most polymorphic bands. AFLP-based genetic distance ranged from 0.0546 to 0.5709 with an average of 0.3799. The dendrogram based on AFLP markers indicated that sweetpotato varieties coming from the same regions or having same parents were clustered in the same groups. Analysis of molecular variance （AMOVA） revealed greater variations within regions （94.08%） than among regions （5.92%）. Thus, the genetic variations mainly existed within regions, while the variations among regions were very low in the tested sweetpotato varieties. Significant genetic variations existed between ＂Northern＂ and ＂Southern＂ sweetpotato varieties when Yangtze River was used as the dividing line.

关键词： AFLP fingerprinting genetic diversity main variety sweetpotato (lpomoea batatas (L.) Lam.)

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The regulatory network mediated by circadian clock genes is related to heterosis in rice

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Journal of Integrative Plant Biology 2015年第3期57卷 300-312页

作者： Guojing Shen Wei Hu Bo Zhang Yongzhong Xing National key laboratory of crop genetic improvement and National Center of Plant Gene Research (Wuhan) Huazhong Agricultural University

Exploitation of heterosis in rice（Oryza sativa L.） has contributed greatly to global food *** this study,we generated three sets of reciprocal F1 hybrids of indica and japonica subspecies to evaluate the relationship between yield heterosis and the circadian *** were no differences in trait performance or heterosis between the reciprocal hybrids,indicating no maternal effects on *** indica-indica and indica-japonica reciprocal F1 hybrids exhibited pronounced heterosis for chlorophyll and starch content in leaves and for grain yield/*** contrast,the japonica-japonica F1 hybrids showed low *** three circadian clock genes investigated expressed in an above-high-parent pattern（AHP）at seedling stage in all the *** five genes downstream of the circadian clock,and involved in chlorophyll and starch metabolic pathways,were expressed in AHP in hybrids with strong better-parent heterosis（BPH）.Similarly,three of these Research Arfive genes in the japonica-japonica F1 hybrids showing low BPH were expressed in positive overdominance,but the other two genes were expressed in additive or negative *** results indicated that the expression patterns of circadian clock genes and their downstream genes are associated with heterosis,which suggests that the circadian rhythm pathway may be related to heterosis in rice.

关键词： Circadian clock better-parent heterosis biomass overdominance yield

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