Recently, two important yield quantitative trait loci (QTLs), GS3 and DEP1, have been cloned in rice (Oryza sativa). Although their relationship has not been established in the ‘rice literature’, a ...recent report identified them as heterotrimeric G protein γ subunits. This identification has profound consequences for our current understanding of both QTLs and the plant G protein signaling network and this opinion article discusses how manipulation of G protein signaling may lead to yield improvements in rice and other crop species. Finally, a mechanistic model to explain the seemingly conflicting phenotypes produced by different GS3 and DEP1 alleles is proposed.
Dear Editor, Rice （Oryza sativa） is the staple food for more than half of the world＇s population. Technologies enabling precise and efficient DNA knock-in or replacement, hereinafter referred to as ...KI, have the potential to revolutionize the generation of crops by precision molecular breeding.
A number of isothermal DNA amplification technologies claim to be ideal for point-of-need (PON) applications as they enable reactions to be performed using a single-temperature heat source (e.g. ...water bath). Thus, we examined several isothermal amplification methods focusing on simplicity, cost, sensitivity and reproducibility to identify the most suitable method(s) for low resource PON applications. A number of methods were found unsuitable as they either involved multiple temperature incubations, were relatively expensive or required relatively large amounts target DNA for amplification. Among the methods examined, loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) were found to be the most suitable for PON applications as they are both single step methods that provide highly sensitive and reproducible amplifications. The speed of LAMP reactions was greatly enhanced, up to 76%, with the addition of loop primers while the presence of swarm primers and the sequestration of free magnesium ions with nucleotides also enhanced the amplification speed. In contrast, we were unable to enhance RPA's performance from the original published literature. While both RPA and LAMP have some drawbacks, either isothermal technology can reliably be used for on-site diagnostics with minimal equipment.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Summary
In rice, amylose content (AC) is controlled by a single dominant Waxy gene. We used Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR‐associated 9 (Cas9) to introduce ...a loss‐of‐function mutation into the Waxy gene in two widely cultivated elite japonica varieties. Our results show that mutations in the Waxy gene reduce AC and convert the rice into glutinous ones without affecting other desirable agronomic traits, offering an effective and easy strategy to improve glutinosity in elite varieties. Importantly, we successfully removed the transgenes from the progeny. Our study provides an example of generating improved crops with potential for commercialization, by editing a gene of interest directly in elite crop varieties.
In this study, we reported generation of new glutinous rice with potential for commercialization by editing Waxy gene in elite crop varieties.
In fungi and metazoans, extracellular signals are often perceived by G-protein-coupled receptors (GPCRs) and transduced through heterotrimeric G-protein complexes to downstream targets. Plant ...heterotrimeric G proteins are also involved in diverse biological processes, but little is known about their upstream receptors. Moreover, the presence of bona fide GPCRs in plants is yet to be established. In Arabidopsis (Arabidopsis thaliana), heterotrimeric G protein consists of one Gα subunit (G PROTEIN α-SUBUNIT1), one Gß subunit (ARABIDOPSIS G PROTEIN ß-SUBUNIT1 AGB1), and three Gys subunits (ARABIDOPSIS G PROTEIN γ-SUBUNIT1 AGG1, AGG2, and AGG3). We identified AGB1 from a suppressor screen of BAK1-interacting receptor-like kinase1-1 (bir1-1), a mutant that activates cell death and defense responses mediated by the receptor-like kinase (RLK) SUPPRESSOR OF BIR1-1. Mutations in AGB1 suppress the cell death and defense responses in bir1-1 and transgenic plants overexpressing SUPPRESSOR OF BIR1-1. In addition, agb1 mutant plants were severely compromised in immunity mediated by three other RLKs, FLAGELLIN-SENSITIVE2 (FLS2), Elongation Factor-TU RECEPTOR (EFR), and CHITIN ELICITOR RECEPTOR KINASE1 (CERK1), respectively. By contrast, G PROTEIN a-SUBUMT1 is not required for either cell death in bir1-1 or pathogen-associated molecular pattern-triggered immunity mediated by FLS2, EFR, and CERK1. Further analysis of agg1 and agg2 mutant plants indicates that AGG1 and AGG2 are also required for pathogen-associated molecular pattern-triggered immune responses mediated by FLS2, EFR, and CERK1, as well as cell death and defense responses in bir1-1. We hypothesize that the Arabidopsis heterotrimeric G proteins function as a converging point of plant defense signaling by mediating responses initiated by multiple RLKs, which may fulfill equivalent roles to GPCRs in fungi and animals.
Heterotrimeric G proteinshave been previously linked to plant defense; however a role for the Gbetagamma dimer in defense signaling has not been described to date. Using available Arabidopsis ...(Arabidopsis thaliana) mutants lacking functional Galpha or Gbeta subunits, we show that defense against the necrotrophic pathogens Alternaria brassicicola and Fusarium oxysporum is impaired in Gbeta-deficient mutants while Galpha-deficient mutants show slightly increased resistance compared to wild-type Columbia ecotype plants. In contrast, responses to virulent (DC3000) and avirulent (JL1065) strains of Pseudomonas syringae appear to be independent of heterotrimeric G proteins. The induction of a number of defense-related genes in Gbeta-deficient mutants were severely reduced in response to A. brassicicola infection. In addition, Gbeta-deficient mutants exhibit decreased sensitivity to a number of methyl jasmonate-induced responses such as induction of the plant defensin gene PDF1.2, inhibition of root elongation, seed germination, and growth of plants in sublethal concentrations of methyl jasmonate. In all cases, the behavior of the Galpha-deficient mutants is coherent with the classic heterotrimeric mechanism of action, indicating that jasmonic acid signaling is influenced by the Gbetagamma functional subunit but not by Galpha. We hypothesize that Gbetagamma acts as a direct or indirect enhancer of the jasmonate signaling pathway in plants.
Nucleic acid amplification is a powerful molecular biology tool, although its use outside the modern laboratory environment is limited due to the relatively cumbersome methods required to extract ...nucleic acids from biological samples. To address this issue, we investigated a variety of materials for their suitability for nucleic acid capture and purification. We report here that untreated cellulose-based paper can rapidly capture nucleic acids within seconds and retain them during a single washing step, while contaminants present in complex biological samples are quickly removed. Building on this knowledge, we have successfully created an equipment-free nucleic acid extraction dipstick methodology that can obtain amplification-ready DNA and RNA from plants, animals, and microbes from difficult biological samples such as blood and leaves from adult trees in less than 30 seconds. The simplicity and speed of this method as well as the low cost and availability of suitable materials (e.g., common paper towelling), means that nucleic acid extraction is now more accessible and affordable for researchers and the broader community. Furthermore, when combined with recent advancements in isothermal amplification and naked eye DNA visualization techniques, the dipstick extraction technology makes performing molecular diagnostic assays achievable in limited resource settings including university and high school classrooms, field-based environments, and developing countries.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The recently developed CRISPR/Cas9 system is a promising technology for targeted genome editing in a variety of species including plants. However, the first generation systems were designed to target ...one or two gene loci at a time. We designed a new multiplex CRISPR/Cas9 system that allows the co-expression of six sgRNA modules in one binary vector using a simple (three steps) cloning strategy in Arabidopsis. The transcription of the sgRNA modules is under the control of three different RNA Polymerase III-dependent promoters. We tested the efficiency of the new multiplex system by targeting six of the fourteen
PYL
families of ABA receptor genes in a single transformation experiment. One line with mutations in all six targeted
PYLs
was identified from 15 T1 plants. The mutagenesis frequency for the six individual
PYL
targets in the T1 lines ranged from 13 to 93 %. In the presence of ABA, the transgenic line identified as containing mutations in all six
PYL
genes produced the highest germination rate in the T2 progeny (37 %). Among these germinated seedlings, half of the analyzed plants (15/30) were homozygous mutants for at least four targeted genes and two plants (6.7 %) contained homozygous mutations in five of the targeted
PYLs
and the other targeted
PYL
had biallelic mutations. Homozygous sextuple mutants were identified in the T3 progeny and characterized together with previously described triple and sextuple
PYL
mutants. We anticipate that the application of this multiplex CRISPR/Cas9 system will strongly facilitate functional analysis of genes pathways and families.
Most conventional and modern crop-improvement methods exploit natural or artificially induced genetic variations and require laborious characterization of the progenies of multiple generations ...derived from time-consuming genetic crosses. Genome-editing systems, in contrast, provide the means to rapidly modify genomes in a precise and predictable way, making it possible to introduce improvements directly into elite varieties. Here, we describe the range of applications available to agricultural researchers using existing genome-editing tools. In addition to providing examples of genome-editing applications in crop breeding, we discuss the technical and social challenges faced by breeders using genome-editing tools for crop improvement.
Most conventional and modern crop-improvement methods exploit natural or artificially induced genetic variations and require laborious characterization of the progeny of multiple generations of time-consuming genetic crosses. Genome-editing systems, in contrast, provide the means to rapidly modify genomes in a precise and predictable way, making it possible to introduce improvements directly into elite varieties. Here, we describe the range of applications available to agricultural researchers using existing genome-editing tools. In addition to providing examples of genome-editing applications in crop breeding, we discuss the technical and social challenges faced by breeders using genome-editing tools for crop improvement.
Genome editing for plant research and crop improvement Zhan, Xiangqiang; Lu, Yuming; Zhu, Jian‐Kang ...
Journal of integrative plant biology,
January 2021, 2021-Jan, 2021-01-00, 20210101, Letnik:
63, Številka:
1
Journal Article
Recenzirano
Odprti dostop
The advent of clustered regularly interspaced short palindromic repeat (CRISPR) has had a profound impact on plant biology, and crop improvement. In this review, we summarize the state‐of‐the‐art ...development of CRISPR technologies and their applications in plants, from the initial introduction of random small indel (insertion or deletion) mutations at target genomic loci to precision editing such as base editing, prime editing and gene targeting. We describe advances in the use of class 2, types II, V, and VI systems for gene disruption as well as for precise sequence alterations, gene transcription, and epigenome control.
This review summarizes the state‐of‐the‐art developments in CRISPR technologies and their applications in plant biology and crop improvement, from the initial introduction of random small indel mutations at target genomic loci to precision editing such as base editing, prime editing and gene targeting.
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