Effective RNAi-Mediated Silencing of the Mismatch Repair MSH2 Gene Induces Sterility of Tomato Plants but Not an Increase in Meiotic Recombination

In plant breeding, the ability to manipulate meiotic recombination aids in the efficient construction of new allelic compositions of chromosomes and facilitates gene transfer from wild relatives of crop plants. The DNA mismatch repair system antagonizes meiotic recombination. In this research, a trial was conducted to evaluate transgenic tomato plants carrying an RNA interference (RNAi) construct designed to inhibit the expression of the mismatch repair MSH2 gene. To drive the RNAi construct, we used either a pro-SmAMP2 promoter from Stellaria media ANTIMICROBIAL PEPTIDE2 or a Cauliflower mosaic virus 35S promoter (CaMV35S). The results of real-time PCR showed that, with a 16 h light/8 h dark photoperiod, MSH2-RNAi tomato transgenic plants exhibited MSH2 gene transcript contents ranging from 0% to 3% in the leaves, relative to untransformed controls. However, with this lighting mode, the MSH2-RNAi transgenic plants grew slowly, flowered poorly, and did not form seed sets. During cultivation with a 12 h light/12 h dark photoperiod, MSH2-RNAi transgenic plants exhibited MSH2 gene transcript contents ranging from 3% to 42%, relative to untransformed controls. Under these conditions, F1 hybrid seed sets formed for most of the MSH2-RNAi transgenic plants with the RNAi construct driven by the CaMV35S promoter, and for one transformant with the RNAi construct driven by the pro-SmAMP2 promoter. Under conditions of a 12 h light/12 h dark photoperiod, most of the F1 transgenic hybrids showed MSH2 gene transcript contents ranging from 3% to 34% and formed F2 offspring sets, which made it possible to assess the meiotic recombination frequency. We showed that the effective inhibition of MSH2 in MSH2-RNAi tomato transgenic plants is not associated with an increase in meiotic recombination compared to the control, but it stimulates the sterility of plants. It was established that the expression of the MSH2 gene in tomato plants is about 50 times higher with a 12 h light/12 h dark than with a 16 h light/8 h dark photoperiod. It is discussed that, in Solanum lycopersicum tomato plants, which are not sensitive to the day length for flowering, changing the lighting time may be a means of controlling the meiotic recombination frequency within certain limits.

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Poorly Repaired Mismatches in Heteroduplex DNA are Hyper-Recombinagenic in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/142.2.407 ◽

1996 ◽

Vol 142 (2) ◽

pp. 407-416 ◽

Cited By ~ 3

Author(s):

P Manivasakam ◽

Susan M Rosenberg ◽

P J Hastings

Keyword(s):

Saccharomyces Cerevisiae ◽

Mismatch Repair ◽

Genetic Markers ◽

Meiotic Recombination ◽

Repair System ◽

Normal Allele ◽

Heteroduplex Dna ◽

Mismatch Repair System ◽

Postmeiotic Segregation ◽

System Operating

Abstract In yeast meiotic recombination, alleles used as genetic markers fall into two classes as regards their fate when incorporated into heteroduplex DNA. Normal alleles are those that form heteroduplexes that are nearly always recognized and corrected by the mismatch repair system operating in meiosis. High PMS (postmeiotic segregation) alleles form heteroduplexes that are inefficiently mismatch repaired. We report that placing any of several high PMS alleles very close to normal alleles causes hyperrecombination between these markers. We propose that this hyperrecombination is caused by the high PMS allele blocking a mismatch repair tract initiated from the normal allele, thus preventing corepair of the two alleles, which would prevent formation of recombinants. The results of three point crosses involving two PMS alleles and a normal allele suggest that high PMS alleles placed between two alleles that are normally corepaired block that corepair.

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A possible causal relationship between iron deficiency, inhibition of DNA synthesis and reduction of meiotic recombination frequency in Neurospora crassa

MGG Molecular & General Genetics ◽

10.1007/bf00269130 ◽

1972 ◽

Vol 119 (2) ◽

pp. 103-117 ◽

Cited By ~ 5

Author(s):

Lars Landner

Keyword(s):

Iron Deficiency ◽

Dna Synthesis ◽

Neurospora Crassa ◽

Causal Relationship ◽

Meiotic Recombination ◽

Recombination Frequency ◽

Inhibition Of Dna Synthesis

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Construction of CRISPR/Cas9 expression vectors habouring gRNA targeted on SLIAA9 gene of tomato

Vietnam Journal of Biotechnology ◽

10.15625/1811-4989/18/1/15273 ◽

2020 ◽

Vol 18 (1) ◽

pp. 147-156

Author(s):

Bui Manh Minh ◽

Ha Hong Hanh ◽

Le Thi Thu Hien ◽

Huynh Thi Thu Hue

Keyword(s):

Transgenic Plants ◽

Genome Editing ◽

Tomato Fruit ◽

Expression System ◽

Secondary Compounds ◽

Expression Vectors ◽

Tomato Plants ◽

Normal Morphology ◽

Transgenic Tomato Plants ◽

Pcr Test

Tomato (Solanum lycopersicum) is a nutritious fruit containing many secondary compounds with health benefits. The formation of tomato fruit through fertilization is controlled by auxin through Aux/IAA9 and ARF8 proteins. The mutated SlIAA9 gene leads to the parthenocarpic development of fruit or seedless tomato fruit. Nowadays, the CRISPR/Cas9 genome editing system is becoming increasingly popular in modifying desired genes on plant objects. In this study, gRNAs which target on tomato SlIAA9 gene were designed and inserted into CRISPR/Cas9 vectors. In addition, two strains of A. tumefaciens harboring pRGEB31-IAA9G2 and pRGEB32-IAA9G2 vectors carrying CRISPR/Cas9 expression system towards SlIAA9 gene in tomato were successfully created. The strain of A. tumefaciens harboring pRGEB31- IAA9G2 plasmid was used to develop transgenic tomato plants from Micro-Tom variety. PCR test showed that 5/14 plants had the presence of Cas9 gene in T0 plants. The transgenic plants have a normal morphology in comparation with the controls. The evaluation of mutant efficiency, type, and stability of mutations on the SlIAA9 will be conducted on next-generation plants when the mutations are stable and segregated into descendents.

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Silencing cathepsin L expression reduces Myzus persicae protein content and the nutritional value as prey for Coccinella septempunctata

10.1101/451211 ◽

2018 ◽

Author(s):

Imran Rauf ◽

Muhammad Asif ◽

Imran Amin ◽

Rubab Zahra Naqvi ◽

Noroza Umer ◽

...

Keyword(s):

Rna Interference ◽

Transgenic Plants ◽

Protein Content ◽

Pest Control ◽

Myzus Persicae ◽

Nutritive Value ◽

Cathepsin L ◽

Coccinella Septempunctata ◽

List Type ◽

Rnai Construct

AbstractGut-expressed aphid genes, which may be more easily inhibited by RNA interference (RNAi) constructs, are attractive targets for pest control efforts involving transgenic plants. Here we show that expression of cathepsin L, a cysteine protease that functions in aphid guts, can be reduced by expression of an RNAi construct in transgenic tobacco. The effectiveness of this approach is demonstrated by up to 80% adult mortality, reduced fecundity, and delayed nymph production of Myzus persicae (green peach aphids) when cathepsin L expression was reduced by plant-mediated RNAi. Consistent with the function of cathepsin L as a gut protease, M. persicae fed on the RNAi plants had a lower protein content in their bodies and excreted more protein in their honeydew. Larvae of Coccinella septempunctata (seven-spotted ladybugs) grew more slowly on aphids having reduced cathepsin L expression, suggesting that prey insect nutritive value, and not just direct negative effects of the RNAi construct, needs to be considered when producing transgenic plants for RNAi-mediated pest control.HighlightsSilencing expression of cathepsin L by RNA interference reduces protein content of Myzus persicae (green peach aphid) bodies.Honeydew of aphids with cathepsin L silenced contains elevated protein.Cathepsin L is required for efficient protein uptake from phloem sap.Aphids with cathepsin L expression silenced have increased mortality and fewer offspring.Coccinella septempunctata (seven-spotted ladybugs) grow more slowly on aphids with expression of cathepsin L silenced.

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