scholarly journals Establishment of A Rapid and Stable Infected System by Agrobacterium-Mediated Transformation of Germination Seeds in Diploid Strawberry.

2020 ◽  
Author(s):  
Peng-Hao Xu ◽  
Xiao-Jiao Gu ◽  
Yuan-Yue Shen
Keyword(s):  
Reporter Gene ◽  
Gene Function ◽  
Agronomic Traits ◽  
Chlorophyll Synthesis ◽  
Infection Frequency ◽  
Magnesium Chelatase ◽  
Climacteric Fruit ◽  
Vector Construction ◽  
Successful Infection ◽  
Diploid Strawberry

Abstract Background Strawberry (Fragaria) is regarded as a model plant for both Rosaceae and non-climacteric fruit ripening. Although much progress has been made in identification of gene function using stable and transient genetic transformation systems in strawberry, the limitation is, more or less, are present. To this end, development of a rapid, efficient, and stable transformation system is required for strawberry research and breeding. Results Here, using diploid Hawaii-4 (Fragaria vesca) seeds and a reporter gene of CHLH (the H subunit of magnesium chelatase magnesium chelatase) key to chlorophyll synthesis, we first develop a rapid, efficient, and stable infected system by the Agrobacterium-mediated seed infection to silence the reporter gene, reaching an infection frequency with 28.3% through a series of optimization elements, including seed full imbibition and initial germination, shaking infection for 24 h, dark cultivation on MS medium for 3 d at 24 ℃, light culture on MS-Tim medium for 1 week at 24 ℃, and vector construction tagged with fluorescence label. Taken together, radicle-emergence germination seeds, appropriate Agrobacterium concentration and infection time are critical for successful infection, finally obtaining the infected kanamycin-resistant seedlings of T1 generation by infected wild seeds within 1 month and T2 generation-infected plants within 4 months. Conclusions The Agrobacterium-mediated infection of germinating seeds (AMTGS) in diploid strawberry (F. vesca) is first established, providing a useful tool for gene function identification and improved agronomic traits in strawberry.

scholarly journals Establishment of a Rapid and Stable Transgenic System by Agrobacterium-Mediated Transformation of the Germination Seeds in Diploid Strawberry.

2020 ◽  
Author(s):  
Xiao-Jiao Gu ◽  
Yuanyue Shen
Keyword(s):  
Reporter Gene ◽  
Fruit Ripening ◽  
Magnesium Chelatase ◽  
Climacteric Fruit ◽  
Vector Construction ◽  
Radicle Emergence ◽  
Strawberry Breeding ◽  
Transformation Systems ◽  
Germinating Seeds ◽  
Diploid Strawberry

Abstract BackgroundStrawberry (Fragaria) is regarded as a model plant for both Rosaceae and non-climacteric fruit ripening. Although much progress has been made in the identification of gene function using traditional, stable and transient genetic transformation systems in strawberry, the limitation is, more or less, present. Thus, development of a rapid, efficient, and stable transformation system is required for strawberry research and breeding.ResultsHere, using diploid Hawaii-4 (Fragaria vesca) seeds and a reporter gene of CHLH (the H subunit of magnesium chelatase magnesium chelatase) , we first develop a new, rapid, efficient, and stable transgenic system by the Agrobacterium-mediated seed transformation to silence the reporter gene, obtaining a transformation frequency with 10 % through a series of optimization conditions, including full imbibition and initial germination, shaking infection for 24 h, dark cultivation on MS medium for 3 d at 24 ℃, light culture on MS-Tim medium for 1 week at 24 ℃, and vector construction carried fluorescence label. Taken together, radicle-emergence germination seeds, appropriate Agrobacterium concentration and infection time are critical for successful transformation, obtaining transgenic kanamycin-resistant seedlings within 1 month and T2 generation transgenic plants within 4 months. ConclusionsWe first have successfully established Agrobacterium-mediated transformation of germinating seeds (AMTGS) in diploid strawberry (F. vesca), providing a useful tool for studying non-climacteric fruit ripening and strawberry breeding.

A study of leaf-senescence genes in rice based on a combination of genomics, proteomics and bioinformatics

2020 ◽  
Author(s):  
Erhui Xiong ◽  
Zhiyong Li ◽  
Chen Zhang ◽  
Jing Zhang ◽  
Ye Liu ◽  
...  
Keyword(s):  
Leaf Senescence ◽  
Molecular Mechanisms ◽  
Agronomic Traits ◽  
Chlorophyll Synthesis ◽  
Research Progress ◽  
Rice Leaf ◽  
Acid Pathway ◽  
Relationship Of ◽  
The Relationship ◽  
Synthesis And Degradation

Abstract Leaf senescence is a highly complex, genetically regulated and well-ordered process with multiple layers and pathways. Delaying leaf senescence would help increase grain yields in rice. Over the past 15 years, more than 100 rice leaf-senescence genes have been cloned, greatly improving the understanding of leaf senescence in rice. Systematically elucidating the molecular mechanisms underlying leaf senescence will provide breeders with new tools/options for improving many important agronomic traits. In this study, we summarized recent reports on 125 rice leaf-senescence genes, providing an overview of the research progress in this field by analyzing the subcellular localizations, molecular functions and the relationship of them. These data showed that chlorophyll synthesis and degradation, chloroplast development, abscisic acid pathway, jasmonic acid pathway, nitrogen assimilation and ROS play an important role in regulating the leaf senescence in rice. Furthermore, we predicted and analyzed the proteins that interact with leaf-senescence proteins and achieved a more profound understanding of the molecular principles underlying the regulatory mechanisms by which leaf senescence occurs, thus providing new insights for future investigations of leaf senescence in rice.

scholarly journals Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops

2020 ◽  
Vol 21 (3) ◽  
pp. 1024 ◽  
Author(s):  
Florian Veillet ◽  
Laura Perrot ◽  
Anouchka Guyon-Debast ◽  
Marie-Paule Kermarrec ◽  
Laura Chauvin ◽  
...  
Keyword(s):  
Plant Breeding ◽  
Genome Editing ◽  
Gene Function ◽  
Physcomitrella Patens ◽  
Agronomic Traits ◽  
Function Analysis ◽  
Single Amino Acid ◽  
Base Substitution ◽  
Base Editing ◽  
Gene Function Analysis

Genome editing has become a major tool for both functional studies and plant breeding in several species. Besides generating knockouts through the classical CRISPR-Cas9 system, recent development of CRISPR base editing holds great and exciting opportunities for the production of gain-of-function mutants. The PAM requirement is a strong limitation for CRISPR technologies such as base editing, because the base substitution mainly occurs in a small edition window. As precise single amino-acid substitution can be responsible for functions associated to some domains or agronomic traits, development of Cas9 variants with relaxed PAM recognition is of upmost importance for gene function analysis and plant breeding. Recently, the SpCas9-NG variant that recognizes the NGN PAM has been successfully tested in plants, mainly in monocotyledon species. In this work, we studied the efficiency of SpCas9-NG in the model moss Physcomitrella patens and two Solanaceae crops (Solanum lycopersicum and Solanum tuberosum) for both classical CRISPR-generated gene knock-out and cytosine base editing. We showed that the SpCas9-NG greatly expands the scope of genome editing by allowing the targeting of non-canonical NGT and NGA PAMs. The CRISPR toolbox developed in our study opens up new gene function analysis and plant breeding perspectives for model and crop plants.

scholarly journals Fine mapping of the QTL cqSPDA2 for chlorophyll content in Brassica napus L.

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Jingxiu Ye ◽  
Haidong Liu ◽  
Zhi Zhao ◽  
Liang Xu ◽  
Kaixiang Li ◽  
...  
Keyword(s):  
Brassica Napus ◽  
Chlorophyll Content ◽  
Fine Mapping ◽  
Agronomic Traits ◽  
Economic Value ◽  
Chlorophyll Synthesis ◽  
Brassica Napus L ◽  
Oil Crops ◽  
Initial Location ◽  
Dh Lines

Abstract Background Chlorophyll is the most important factor enabling plants to absorb, transfer and transform light energy and plays an important role in yield formation. Brassica napus is one of the most important oil crops. Breeding Brassica napus for high light efficiency by improving photosynthetic efficiency has considerable social and economic value. In Brassica napus, there have been studies of the initial location of chlorophyll in seed embryos and pericarps, but there are few reports on the fine mapping of chlorophyll QTLs. We constructed near-isogenic lines (NIL), fine-mapped a chlorophyll locus, and evaluated the effect of this dominant locus on agronomic traits. Results The cqSPDA2 locus was mapped to an interval of 21.87–22.91 Mb on the chromosome A02 of Brassica napus using doubled haploid (DH) lines. To fine-map cqSPDA2, we built NIL and designed Indel primers covering the mapping interval. The 469 individuals in the BC3F2 population were analyzed using these indel primers. Among these indel primers, 15 could narrow the mapping interval to 188 kb between Indel3 and Indel15. Next, 16 indel primers and 19 SSR primers were designed within the new narrower mapping interval, and 5 of the primer-amplified fragments were found to be polymorphic and tightly linked to the cqSPDA2 locus in the BC4F2 population. The mapping interval was narrowed to 152 kb on A02 between SSR2 and Indel15. By gene expression analysis, we found three annotated genes in the mapping interval, including BnaA02g30260D, BnaA02g30290D and BnaA02g30310D, which may be responsible for chlorophyll synthesis. Conclusions The locus cqSPDA2, a dominant QTL for chlorophyll content in Brassica napus, was fine-mapped to a 21.89–22.04 Mb interval on A02. Three annotated genes (BnaA02g30260D, BnaA02g30290D and BnaA02g30310D) that may be responsible for chlorophyll synthesis were found.

1-N-histidine phosphorylation of ChlD by the AAA+ ChlI2 stimulates magnesium chelatase activity in chlorophyll synthesis

2017 ◽  
Vol 474 (12) ◽  
pp. 2095-2105 ◽  
Author(s):  
Artur Sawicki ◽  
Shuaixiang Zhou ◽  
Kathrin Kwiatkowski ◽  
Meizhong Luo ◽  
Robert D. Willows
Keyword(s):  
Rhodobacter Capsulatus ◽  
Protoporphyrin Ix ◽  
Chlorophyll Synthesis ◽  
Magnesium Chelatase ◽  
Orthologous Protein ◽  
Substrate Complex ◽  
Motor Complex ◽  
Histidine Phosphorylation ◽  
Mg Chelatase ◽  
Stimulation Of

Magnesium chelatase (Mg-chelatase) inserts magnesium into protoporphyrin during the biosynthesis of chlorophyll and bacteriochlorophyll. Enzyme activity is reconstituted by forming two separate preactivated complexes consisting of a GUN4/ChlH/protoporphyrin IX substrate complex and a ChlI/ChlD enzyme ‘motor’ complex. Formation of the ChlI/ChlD complex in both Chlamydomonas reinhardtii and Oryza sativa is accompanied by phosphorylation of ChlD by ChlI, but the orthologous protein complex from Rhodobacter capsulatus, BchI/BchD, gives no detectable phosphorylation of BchD. Phosphorylation produces a 1-N-phospho-histidine within ChlD. Proteomic analysis indicates that phosphorylation occurs at a conserved His residue in the C-terminal integrin I domain of ChlD. Comparative analysis of the ChlD phosphorylation with enzyme activities of various ChlI/ChlD complexes correlates the phosphorylation by ChlI2 with stimulation of Mg-chelatase activity. Mutation of the H641 of CrChlD to E641 prevents both phosphorylation and stimulation of Mg-chelatase activity, confirming that phosphorylation at H641 stimulates Mg-chelatase. The properties of ChlI2 compared with ChlI1 of Chlamydomonas and with ChlI of Oryza, shows that ChlI2 has a regulatory role in Chlamydomonas.

scholarly journals C-terminal residues of Oryza sativa GUN4 are required for the activation of the ChlH subunit of magnesium chelatase in chlorophyll synthesis

FEBS Letters ◽  
2012 ◽  
Vol 586 (3) ◽  
pp. 205-210 ◽  
Author(s):  
Shuaixiang Zhou ◽  
Artur Sawicki ◽  
Robert D. Willows ◽  
Meizhong Luo
Keyword(s):  
Oryza Sativa ◽  
Chlorophyll Synthesis ◽  
Magnesium Chelatase

scholarly journals Chlorophyll Biosynthesis. Expression of a Second Chl I Gene of Magnesium Chelatase in Arabidopsis Supports Only Limited Chlorophyll Synthesis

2002 ◽  
Vol 128 (2) ◽  
pp. 770-779 ◽  
Author(s):  
Heather M. Rissler ◽  
Eva Collakova ◽  
Dean DellaPenna ◽  
James Whelan ◽  
Barry J. Pogson
Keyword(s):  
Chlorophyll Biosynthesis ◽  
Chlorophyll Synthesis ◽  
Magnesium Chelatase ◽  
I Gene

scholarly journals Fine mapping of the QTL cqSPDA2 for chlorophyll content in Brassica napus L.

2020 ◽  
Author(s):  
Jingxiu Ye ◽  
Haidong Liu ◽  
Zhi Zhao ◽  
Liang Xu ◽  
Kaixiang Li ◽  
...  
Keyword(s):  
Brassica Napus ◽  
Chlorophyll Content ◽  
Fine Mapping ◽  
Agronomic Traits ◽  
Economic Value ◽  
Chlorophyll Synthesis ◽  
Brassica Napus L ◽  
Oil Crops ◽  
Initial Location ◽  
Dh Lines

Abstract Background: Chlorophyll is the most important factor enabling plants to absorb, transfer and transform light energy and plays an important role in yield formation. Brassica napus is one of the most important oil crops. Breeding Brassica napus for high light efficiency by improving photosynthetic efficiency has considerable social and economic value. In Brassica napus, there have been studies of the initial location of chlorophyll in seed embryos and pericarps, but there are few reports on the fine mapping of chlorophyll QTLs. We constructed near-isogenic lines (NIL), fine-mapped a chlorophyll locus, and evaluated the effect of this dominant locus on agronomic traits.Results: The cqSPDA2 locus was mapped to an interval of 21.87-22.91 Mb on the chromosome A02 of Brassica napus using doubled haploid (DH) lines. To fine-map cqSPDA2, we built NIL and designed Indel primers covering the mapping interval. The 469 individuals in the BC3F2 population were analyzed using these indel primers. Among the indel primers, 15 could narrow the mapping interval to 188 kb between Indel3 and Indel15. Next, 16 indel primers and 19 SSR primers were designed within the new narrower mapping interval, and 5 of the primer-amplified fragments were found to be polymorphic and tightly linked to the cqSPDA2 locus in the BC4F2 population. The mapping interval was narrowed to 152 kb on A02 between SSR2 and Indel15. By gene expression analysis, we found three annotated genes in the mapping interval, including BnaA02g30260D, BnaA02g30290D and BnaA02g30310D, which may be responsible for chlorophyll synthesis. Conclusions: The locus cqSPDA2, a dominant QTL for chlorophyll content in Brassica napus, was fine-mapped to a 21.89-22.04 Mb interval on A02. Three annotated genes (BnaA02g30260D, BnaA02g30290D and BnaA02g30310D) that may be responsible for chlorophyll synthesis were found.

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