PE6c greatly enhances prime editing in transgenic rice plants.

Prime editing is a versatile CRISPR/Cas-based precise genome-editing technique for crop breeding. Four new types of prime editors (PEs) named PE6a-d were recently generated using evolved and engineered reverse transcriptase (RT) variants from three different sources. In this study, we tested the editing efficiencies of four PE6 variants and two additional PE6 constructs with double-RT modules in transgenic rice (Oryza sativa) plants. PE6c, with an evolved and engineered RT variant from the yeast Tf1 retrotransposon, yielded the highest prime-editing efficiency. The average fold change in the editing efficiency of PE6c compared with PEmax exceeded 3.5 across 18 agronomically important target sites from 15 genes. We also demonstrated the feasibility of using two RT modules to improve prime-editing efficiency. Our results suggest that PE6c or its derivatives would be an excellent choice for prime editing in monocot plants. In addition, our findings have laid a foundation for prime-editing-based breeding of rice varieties with enhanced agronomically important traits.

High-efficiency prime editing enables new strategies for broad-spectrum resistance to bacterial blight of rice.

Using genetic resistance against bacterial blight (BB) caused by Xanthomonas oryzae pathovar oryzae (Xoo) is a major objective in rice breeding programmes. Prime editing (PE) has the potential to create novel germplasm against Xoo. Here, we use an improved prime-editing system to implement two new strategies for BB resistance. Knock-in of TAL effector binding elements (EBE) derived from the BB susceptible gene SWEET14 into the promoter of a dysfunctional executor R gene xa23 reaches 47.2% with desired edits including biallelic editing at 18% in T0 generation that enables an inducible TALE-dependent BB resistance. Editing the transcription factor TFIIA gene TFIIAγ5 required for TAL effector-dependent BB susceptibility recapitulates the resistance of xa5 at an editing efficiency of 88.5% with biallelic editing rate of 30% in T0 generation. The engineered loci provided resistance against multiple Xoo strains in T1 generation. Whole-genome sequencing detected no OsMLH1dn-associated random mutations and no off-target editing demonstrating high specificity of this PE system. This is the first-ever report to use PE system to engineer resistance against biotic stress and to demonstrate knock-in of 30-nucleotides cis-regulatory element at high efficiency. The new strategies hold promises to fend rice off the evolving Xoo strains and protect it from epidemics.

Optimized prime editing efficiently generates heritable mutations in maize.

Low efficiency is the main obstacle to using prime editing in maize (Zea mays). Recently, prime-editing efficiency was greatly improved in mammalian cells and rice (Oryza sativa) plants by engineering prime-editing guide RNAs (pegRNAs), optimizing the prime editor (PE) protein, and manipulating cellular determinants of prime editing. In this study, we tested PEs optimized via these three strategies in maize. We demonstrated that the ePE5max system, composed of PEmax, epegRNAs (pegRNA-evopreQ. 1), nicking single guide RNAs (sgRNAs), and MLH1dn, efficiently generated heritable mutations that conferred resistance to herbicides that inhibit 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), acetolactate synthase (ALS), or acetyl CoA carboxylase (ACCase) activity. Collectively, we demonstrate that the ePE5max system has sufficient efficiency to generate heritable (homozygous or heterozygous) mutations in maize target genes and that the main obstacle to using PEs in maize has thus been removed.

Comprehensive quality evaluation of Polygoni Orientalis Fructus and its processed product: chemical fingerprinting and simultaneous determination of seven major components coupled with chemometric analyses.

INTRODUCTION: Polygoni Orientalis Fructus (POF) is a clinically effective Chinese medicine. Raw POF (RPOF) and POF Tostus (POFT) are used separately in clinics. However, incomplete progress has been made on quality control. OBJECTIVE: To establish a comprehensive method for quality assessment of RPOF and POFT and to discriminate these two varieties. METHODOLOGY: High-performance liquid chromatography combined with the diode array detector (HPLC-DAD) methods were developed for fingerprinting and quantitative analysis of seven major compounds in RPOF and POFT, and the main components were determined by HPLC-DAD coupled with Fourier-transform ion cyclotron resonance-mass spectrometry. Chemometric approaches were performed to discriminate RPOF and POFT and to screen discriminatory components. RESULTS: Fingerprints were established and 12 common peaks were identified, cannabisin G and cannabisin E were firstly identified from POF. In quantitative analysis, all analytes showed good regression (R > 0.9996) within test ranges and the recovery of the method was in the range 96.6-104.3%. Fingerprints in conjunction with similarity analysis and hierarchical clustering analysis (HCA) demonstrated the consistent quality of RPOF and showed a clear discrimination between RPOF and POFT. Principal component analysis, partial least-squares discriminant analysis, and heatmap-HCA on quantitative data not only gave a clear differentiation between RPOF and POFT, but they also suggested that quercetin, 3,5,7-trihydroxychromone, and N-trans-feruloyltyramine acted as the main factors responsible for the sample differences. CONCLUSIONS: Chromatographic analysis in combination with chemometric analysis provides a simple and reliable method of comparing and evaluating the qualities of RPOF and POFT.

A Novel Ternary Vector System United with Morphogenic Genes Enhances CRISPR/Cas Delivery in Maize.

The lack of efficient delivery methods is a major barrier to clustered regularly interspaced short palindromic repeats/CRISPR-associated protein (CRISPR/Cas)-mediated genome editing in many plant species. Combinations of morphogenic regulator (MR) genes and ternary vector systems are promising solutions to this problem. In this study, we first demonstrated that MR vectors greatly enhance maize (Zea mays) transformation. We then tested a CRISPR/Cas9 MR vector in maize and found that the MR and CRISPR/Cas9 modules have no negative influence on each other. Finally, we developed a novel ternary vector system to integrate the MR and CRISPR/Cas modules. Our ternary vector system is composed of new pGreen-like binary vectors, here named pGreen3, and a pVS1-based virulence helper plasmid, which also functions as a replication helper for the pGreen3 vectors in Agrobacterium tumefaciens The pGreen3 vectors were derived from the plasmid pRK2 and display advantages over pGreen2 vectors regarding both compatibility and stability. We demonstrated that the union of our ternary vector system with MR gene modules has additive effects in enhancing maize transformation and that this enhancement is especially evident in the transformation of recalcitrant maize inbred lines. Collectively, our ternary vector system-based tools provide a user-friendly solution to the low efficiency of CRISPR/Cas delivery in maize and represent a basic platform for developing efficient delivery tools to use in other plant species recalcitrant to transformation.

Potential high-frequency off-target mutagenesis induced by CRISPR/Cas9 in Arabidopsis and its prevention.

KEY MESSAGE: We present novel observations of high-specificity SpCas9 variants, sgRNA expression strategies based on mutant sgRNA scaffold and tRNA processing system, and CRISPR/Cas9-mediated T-DNA integrations. Specificity of CRISPR/Cas9 tools has been a major concern along with the reports of their successful applications. We report unexpected observations of high frequency off-target mutagenesis induced by CRISPR/Cas9 in T1 Arabidopsis mutants although the sgRNA was predicted to have a high specificity score. We also present evidence that the off-target effects were further exacerbated in the T2 progeny. To prevent the off-target effects, we tested and optimized two strategies in Arabidopsis, including introduction of a mCherry cassette for a simple and reliable isolation of Cas9-free mutants and the use of highly specific mutant SpCas9 variants. Optimization of the mCherry vectors and subsequent validation found that fusion of tRNA with the mutant rather than the original sgRNA scaffold significantly improves editing efficiency. We then examined the editing efficiency of eight high-specificity SpCas9 variants in combination with the improved tRNA-sgRNA fusion strategy. Our results suggest that highly specific SpCas9 variants require a higher level of expression than their wild-type counterpart to maintain high editing efficiency. Additionally, we demonstrate that T-DNA can be inserted into the cleavage sites of CRISPR/Cas9 targets with high frequency. Altogether, our results suggest that in plants, continuous attention should be paid to off-target effects induced by CRISPR/Cas9 in current and subsequent generations, and that the tools optimized in this report will be useful in improving genome editing efficiency and specificity in plants and other organisms.

A CRISPR/Cas9 toolkit for multiplex genome editing in plants.

BACKGROUND: To accelerate the application of the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/ CRISPR-associated protein 9) system to a variety of plant species, a toolkit with additional plant selectable markers, more gRNA modules, and easier methods for the assembly of one or more gRNA expression cassettes is required. RESULTS: We developed a CRISPR/Cas9 binary vector set based on the pGreen or pCAMBIA backbone, as well as a gRNA (guide RNA) module vector set, as a toolkit for multiplex genome editing in plants. This toolkit requires no restriction enzymes besides BsaI to generate final constructs harboring maize-codon optimized Cas9 and one or more gRNAs with high efficiency in as little as one cloning step. The toolkit was validated using maize protoplasts, transgenic maize lines, and transgenic Arabidopsis lines and was shown to exhibit high efficiency and specificity. More importantly, using this toolkit, targeted mutations of three Arabidopsis genes were detected in transgenic seedlings of the T1 generation. Moreover, the multiple-gene mutations could be inherited by the next generation. CONCLUSIONS: We developed a toolkit that facilitates transient or stable expression of the CRISPR/Cas9 system in a variety of plant species, which will facilitate plant research, as it enables high efficiency generation of mutants bearing multiple gene mutations.

Modularly assembled multiplex prime editors for simultaneous editing of agronomically important genes in rice.

Prime editing (PE) technology enables precise alterations in the genetic code of a genome of interest. PE offers great potential for identifying major agronomically important genes in plants and editing them into superior variants, ideally targeting multiple loci simultaneously to realize the collective effects of the edits. Here, we report the development of a modular assembly-based multiplex PE system in rice and demonstrate its efficacy in editing up to four genes in a single transformation experiment. The duplex PE (DPE) system achieved a co-editing efficiency of 46.1% in the T0 generation, converting TFIIAγ5 to xa5 and xa23 to Xa23SW11. The resulting double-mutant lines exhibited robust broad-spectrum resistance against multiple Xanthomonas oryzae pathovar oryzae (Xoo) strains in the T1 generation. In addition, we successfully edited OsEPSPS1 to an herbicide-tolerant variant and OsSWEET11a to a Xoo-resistant allele, achieving a co-editing rate of 57.14%. Furthermore, with the quadruple PE (QPE) system, we edited four genes-two for herbicide tolerance (OsEPSPS1 and OsALS1) and two for Xoo resistance (TFIIAγ5 and OsSWEET11a)-using one construct, with a co-editing efficiency of 43.5% for all four genes in the T0 generation. We performed multiplex PE using five more constructs, including two for triplex PE (TPE) and three for QPE, each targeting a different set of genes. The editing rates were dependent on the activity of pegRNA and/or ngRNA. For instance, optimization of ngRNA increased the PE rates for one of the targets (OsSPL13) from 0% to 30% but did not improve editing at another target (OsGS2). Overall, our modular assembly-based system yielded high PE rates and streamlined the cloning of PE reagents, making it feasible for more labs to utilize PE for their editing experiments. These findings have significant implications for advancing gene editing techniques in plants and may pave the way for future agricultural applications.

Enhanced editing efficiency in Arabidopsis with a LbCas12a variant harboring D156R and E795L mutations.

Cas12a (Cpf1), a Class 2 Type V CRISPR/Cas nuclease, has several unique attributes for genome editing and may provide a valuable alternative to Cas9. However, a low editing efficiency due to temperature sensitivity and insufficient cleavage activity of the Cas12a nuclease are major obstacles to its broad application. In this report, we generated two variants, ttAsCas12 Ultra and ttLbCas12a Ultra harboring three (E174R, M537R, and F870L) or two (D156R and E795L) mutations, respectively, by combining the mutations from the temperature-tolerant variants ttAsCas12a (E174R) and ttLbCas12a (D156R), and those from the highly active variants AsCas12a Ultra (M537R and F870L) and LbCas12a Ultra (E795L). We compared editing efficiencies of the five resulting Cas12a variants (LbCas12a, ttLbCas12a, ttLbCas12a Ultra, AsCas12a Ultra, and ttAsCas12 Ultra) at six target sites of four genes in Arabidopsis (Arabidopsis thaliana). The variant ttLbCas12a Ultra, harboring the D156R and E795L mutations, exhibited the highest editing efficiency of all variants tested in Arabidopsis and can be used to generate homozygous or biallelic mutants in a single generation in Arabidopsis plants grown at 22 °C. In addition, optimization of ttLbCas12a Ultra, by varying nuclear localization signal sequences and codon usage, further greatly improved editing efficiency. Collectively, our results indicate that ttLbCas12a Ultra is a valuable alternative to Cas9 for editing genes or promoters in Arabidopsis. Supplementary Information: The online version contains supplementary material available at 10.1007/s42994-024-00144-w.

[The binding property of different regions in Duffy-binding-like domain alpha of Plasmodium falciparum erythrocyte membrane protein 1 to heparin].

OBJECTIVE: To clone and express Plasmodium falciparum erythrocyte membrane protein 1 DBLalpha (PfEMP1-DBLalpha) and three fragments genes, and screen the strongest affinity sequence with the red blood cell surface receptors-heparin or heparin sulfate in the structure of PfEMP1-DBLalpha. METHODS: The sequence of PfEMP1-DBLalpha1245 was optimized according to the characteristics of E. coli codon, synthesized, and divided into three fragments (DBLaA, DBLalphaB, and DBLalphaC) by PCR. Full-length gene and three gene fragments were subcloned into PGEX-4T-1 vector, and transformed into E. coli BL21 and then induced with IPTG for expression. The recombinant protein was purified from bacterial lysates using glutathione-Sepharose 4B. Heparin affinity test and glycosaminoglycan (GAG) inhibition test were used to analyze the affinity between recombinant protein and heparin. RESULTS: Four recombinant proteins(DBLalpha1245, DBLalphaA, DBLalphaB, and DBLalphaC) were expressed as solubility and the relative molecular weight (M(r) 73 600, M(r) 41 600, M(r) 42 500, and M(r) 41 500) were conformed to the prediction size. Heparin affinity test and GAG inhibition test showed that the four recombinant proteins were binded to the heparin-Sepharose, but not for the GST control. DBLalphaC (Q285-Y415) had the strongest affinity to heparin. CONCLUSION: The strongest affinity sequence with heparin or heparin sulfate in the structure of PfEMP1-DBLalpha is Q285-Y415, which plays a role in binding of Plasmodium falciparum infected red blood cells to the peripheral red blood cells.

MISSA is a highly efficient in vivo DNA assembly method for plant multiple-gene transformation.

We describe a highly efficient in vivo DNA assembly method, multiple-round in vivo site-specific assembly (MISSA), which facilitates plant multiple-gene transformation. MISSA is based on conjugational transfer, which is driven by donor strains, and two in vivo site-specific recombination events, which are mediated by inducible Cre recombinase and phage lambda site-specific recombination proteins in recipient strains, to enable in vivo transfer and in vivo assembly of multiple transgenic DNA. The assembly reactions can be performed circularly and iteratively through alternate use of the two specially designed donor vectors. As proof-of-principle experiments, we constructed a few plant multigene binary vectors. One of these vectors was generated by 15 rounds of MISSA reactions and was confirmed in transgenic Arabidopsis (Arabidopsis thaliana). As MISSA simplifies the tedious and time-consuming in vitro manipulations to a simple mixing of bacterial strains, it will greatly save time, effort, and expense associated with the assembly of multiple transgenic or synthetic DNA. The principle that underlies MISSA is applicable to engineering polygenic traits, biosynthetic pathways, or protein complexes in all organisms, such as Escherichia coli, yeast, plants, and animals. MISSA also has potential applications in synthetic biology, whether for basic theory or for applied biotechnology, aiming at the assembly of genetic pathways for the production of biofuels, pharmaceuticals, and industrial compounds from natural or synthetic DNA.

[Cloning and expression of var2csa DBL domains from Plasmodium falciparum hainan isolate and functional analysis of the recombinant protein].

OBJECTIVE: To clone and express three VAR2CSA duffy antigen-binding ligand (DBL) domains (DBL4/ 5/6) encoded by var2csa gene of a Hainan isolate of Plasmodium falciparum, and study the difference of chondroitin sulfate A (CSA)-binding activity among them. METHODS: Three DBL domains was amplified by PCR and cloned into the vector pMD18-T. The recombinant plasmids were identified by enzyme digestion and sequencing, and then subcloned into the prokaryotic expression vector pET-22b. The recombinant plasmid was transformed into E. coli BL21 (DE3) and followed by expression of the protein induced by IPTG. The recombinant protein was purified with His GraciTrap kit and identified by SDS-PAGE and Western blotting. CSA-binding activity of the three recombinant DBL domains was assayed by ELISA. RESULTS: The target genes were amplified with the length of 996 bp, 859 bp and 894 bp. The constructed recombinant plasmids were identified by enzyme digestion and DNA sequencing. The recombinant proteins (DBL4/5/6) were purified, the relative molecular mass of DBLfA, DBL5 and DBL6 was Mr 439 800, Mr 34,500 and Mr 36,000, respectively. The purified protein has been confirmed with immunogenicity by Western blotting. The results of adhesion experiment indicated that A405 values of DBL5 domain with different concentration were significantly higher than that of DBLA and DBL6. CONCLUSION: The three recombinant proteins (DBLA/5/6) of VAR2CSA DBL domains were expressed, and DBL5 domain has high binding affinity with CSA.

Efficacy of uremic pruritus treatment in patients undergoing hemodialysis, a network meta-analysis for randomized clinical trials.

INTRODUCTION: Uremic pruritus is very common in hemodialysis or renal failure patients, there were lots of available treatments such as gabapentin, pregabalin, ondansetron, etc. However, there is no quantified study comparing these treatments together, it is impossible to conduct a clinical trial involving so many treatments, so we conduct a network meta-analysis to compare them. METHOD: We collected mean difference and standard error of visual analogue scale data as outcome. In total we collected 15 articles, 15 articles, 1180 subjects and 6 treatments included to this study. RESULTS: In these comparisons, gabapentin showed the largest effect MD: 5.19, 95%CI [3.77, 6.61], anti-histamine MD: 4.65, 95%CI [2.22, 7.07] and pregabalin MD: 4.62, 95%CI [2.71, 6.62] showed a similar effect. Opioid pathway related treatment also showed a significant but not so large effect MD: 2.45, 95%CI [0.41, 4.49]. Ondansetron and Doxepin didn't show a significant improvement among placebo, the overall quantifying heterogeneity I2=43.1%. There is no statically difference between gabapentin, pregabalin and anti-histamine treatments. CONCLUSIONS: So we conclude that gabapentin, pregabalin and anti-histamine has a similar efficacy on pruritus control.

[Polysaccharide and molecular pathogenesis of Plasmodium falciparum].

In the interaction of Plasmodium falciparum with human cells, sporozoite adheres to the receptor of the liver endothelial cell, then invades to liver. Merozoite binds to the surface of red blood cells, and invades to erythrocyte. The adhesion of membrane protein of the infected erythrocytes to the surface molecules of vascular endothelial cell in the vital organs leads to the obstruction of blood circulation eventually. The adhesion is mediated by interaction between parasite-derived ligands and the negative charged polysaccharides on the surface of host cells. This review is to discuss the molecular mechanisms in the host-parasite interactions.

Agrobacterium-mediated delivery of CRISPR/Cas reagents for genome editing in plants enters an era of ternary vector systems.

Lack of appropriate methods for delivery of genome-editing reagents is a major barrier to CRISPR/Cas-mediated genome editing in plants. Agrobacterium-mediated genetic transformation (AMGT) is the preferred method of CRISPR/Cas reagent delivery, and researchers have recently made great improvements to this process. In this article, we review the development of AMGT and AMGT-based delivery of CRISPR/Cas reagents. We give an overview of the development of AMGT vectors including binary vector, superbinary vector, dual binary vector, and ternary vector systems. We also review the progress in Agrobacterium genomics and Agrobacterium genetic engineering for optimal strains. We focus in particular on the ternary vector system and the resources we developed. In summary, it is our opinion that Agrobacterium-mediated CRISPR/Cas genome editing in plants is entering an era of ternary vector systems, which are often integrated with morphogenic regulators. The new vectors described in this article are available from Addgene and/or MolecularCloud for sharing with academic investigators for noncommercial research.

The clinical effect and safety of the treatment of tibia intercondylar eminence fracture with cannulated screw and suture fixation under arthroscope: Protocol for a systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: The clinical effects and safety over the treatment of tibia intercondylar eminence fracture (TIEF) with cannulated screw and suture fixation were evaluated under arthroscope systematically, providing evidence-based medical support for the selection of surgical methods in terms of minimally invasive arthroscopic treatment for TIEF. METHODS: The English databases of PubMed, EMBASE, Cochrane Library, CNKI, SinoMed, VIP, and Wanfang databases were searched by computer. The randomized controlled trials were conducted to compare the clinical effects of TIEF with cannulated screw and suture fixation under arthroscope. The retrieval period is from the beginning of database building to January 2020. There is no language restriction. Chinese databases are searched by keywords, while English databases are searched by the combination of subject words and free words. According to the retrieval strategy, the two evaluators will lead the conforming documents into Note Express for repeated literature screening, and the two evaluators will extract and cross-check the conforming documents according to the pre-designed data extraction table. Two researchers adopted the modified Jadad scale independently to evaluate the quality of the literature. The RevMan 5.3 version software provided by the Cochrane Collaboration Network was adopted for statistical analysis. RESULTS: The study will strictly review and extract the data included in the literature, and scientifically make statistical analysis for the pre-set outcome indicators. All the research processes will be conducted in strict accordance with the guidance of system evaluation. In this study, the differences between cannulated screw fixation and suture fixation under arthroscopy will be evaluated by comparing the relevant outcome indicators. All the results of this study will be published openly in a highly influential professional academic journal. CONCLUSION: The paper adopted Cochrane system evaluation method to collect and sort out the published literature about the treatment of tibial eminence fracture between cannulated screw fixation and suture fixation under arthroscopy, and to compare the clinical efficacy and safety of the two fixation methods utilizing meta-analysis and comparison of related outcome indicators. Through this study, we will draw a positive conclusion, which will provide a basis for the better treatment of tibial eminence fracture. PROSPERO REGISTRATION NUMBER: PROSPERO CRD42020168433.

Prime editing efficiently generates W542L and S621I double mutations in two ALS genes in maize.

Prime editing is a novel and universal CRISPR/Cas-derived precision genome-editing technology that has been recently developed. However, low efficiency of prime editing has been shown in transgenic rice lines. We hypothesize that enhancing pegRNA expression could improve prime-editing efficiency. In this report, we describe two strategies for enhancing pegRNA expression. We construct a prime editing vector harboring two pegRNA variants for W542L and S621I double mutations in ZmALS1 and ZmALS2. Compared with previous reports in rice, we achieve much higher prime-editing efficiency in maize. Our results are inspiring and provide a direction for the optimization of plant prime editors.

The art of microRNA: various strategies leading to gene silencing via an ancient pathway.

MicroRNAs (miRNAs), an endogenous type of small RNAs of approximately 22 nucleotides (nt), have long resided in the cells of plants and animals including humans, constituting an ancient pathway of gene regulation in eukaryotes. They have a simple structure in their mature form but carry enormous information that may regulate up to 90% of the human transcriptome. Furthermore, the multi-facets of a miRNA are tightly associated with diverse cellular proteins that make it broadly connected to various physiological and pathological processes. This review aims to examine miRNAs briefly from their biogenesis to their general functions with an emphasis on working mechanisms in regulation of their target mRNAs.

Differential and dynamic regulation of miR398 in response to ABA and salt stress in Populus tremula and Arabidopsis thaliana.

MicroRNAs (miRNAs) are endogenous small RNAs of ~22 nucleotides (nt) that play a key role in down regulation of gene expression at the post-transcriptional level in plants and animals. Various studies have identified numerous miRNAs that were either up regulated or down regulated upon stress treatment. Here, we sought to understand the temporal regulation of miRNAs in different plant species under abscisic acid (ABA) and salt (NaCl) stress. Our results showed that the regulation of miR398 in response to ABA and salt stress was more dynamic in plants than previously reported. In poplars, miR398 was first induced upon 3-4 h of ABA or salt stress. However, this induction declined after 48 h and finally accumulated again over a prolonged stress (72 h). We referred to this kind of regulation as dynamic regulation. In contrast, such dynamic regulation of miR398 under salt stress was completely absent in Arabidopsis, in which miR398 was steadily and unidirectionally suppressed. Interestingly, ABA treatment caused a deviate dynamic regulation of miR398 in Arabidopsis, showing an opposite response as compared to that in poplars. We referred to the difference in regulation between Arabidopsis and poplars as differential regulation. Furthermore, the expression of the miR398 target, copper superoxide dismutase1 (CSD1), was in reverse correlation with the miR398 level, suggesting a control of this specific target expression predominantly by miR398 under abiotic stress. Together, these data consistently show a correlated regulation between miR398 and its representative target, CSD1, by ABA and salt stresses, and raise the possibility that regulation of miRNAs in plants is twofold: a dynamic regulation within a plant species and a differential regulation between different plant species.

[Screening for relevant proteins involved in adhesion of Cryptosporidium parvum sporozoites to host cells].

The Cryptosporidium parvum T7 phage display library was screened by using Caco-2 cells. Five specific gene fragments were identified by blasting sequences in GenBank, one of which encoding the CP2 protein was previously identified as a surface molecule of sporozoites and involved in parasite invasion. The others are hypothetic proteins with unknown functions. Bioinformatic analysis of these proteins indicated that they may be involved in the host-parasite interactions.