scholarly journals Analysis of amplicons containing the esterase genes responsible for insecticide resistance in the peach-potato aphid Myzus persicae (Sulzer)

1996 ◽  
Vol 313 (2) ◽  
pp. 543-547 ◽  
Author(s):  
Linda M. FIELD ◽  
Alan L. DEVONSHIRE ◽  
Chris TYLER-SMITH
Keyword(s):  
Gene Amplification ◽  
Myzus Persicae ◽  
Gene Copy Number ◽  
Fold Increase ◽  
Gene Copy ◽  
Potato Aphid ◽  
Gene Copies ◽  
Genes Encoding ◽  
Aphid Clone ◽  
Peach Potato Aphid

The amplification of genes encoding an insecticide-detoxifying esterase (E4) in the peach-potato aphid Myzus persicae is one of the few examples where this genetic phenomenon has been shown to be involved in the response of an intact higher organism to artificial selection. Here we report quantitative and qualitative studies of the repeat units (amplicons) containing the E4 genes in a highly resistant aphid clone. Initial studies to quantify esterase sequences showed a 5-11-fold increase in resistant aphids compared with susceptible aphids, suggesting the presence of 10-22 gene copies per diploid genome. A more incisive analysis by pulsed-field gel electrophoresis confirmed the presence of about 12 copies of the E4 gene and showed them to be on about 24 kb amplicons, arranged as a tandem array of direct repeats. This, together with previous results from crossing experiments and with recent in situ hybridization studies, confirms that the E4 gene amplification in this aphid clone is heterozygous at a single locus. However, these data show that the gene amplification alone cannot account for the approx. 60 times higher levels of E4 protein and its mRNA present in this aphid clone, and therefore resistance must involve changes in both esterase gene copy number and gene expression.

scholarly journals Relationship between amount of esterase and gene copy number in insecticide-resistant Myzus persicae (Sulzer)

1999 ◽  
Vol 339 (3) ◽  
pp. 737-742 ◽  
Author(s):  
Linda M. FIELD ◽  
Roger L. BLACKMAN ◽  
Chris TYLER-SMITH ◽  
Alan L. DEVONSHIRE
Keyword(s):  
Gene Amplification ◽  
Myzus Persicae ◽  
Copy Number ◽  
Transcriptional Control ◽  
Gene Copy Number ◽  
Gene Copy ◽  
Copy Numbers ◽  
And Function ◽  
Gene Copy Numbers

Overproduction of the insecticide-degrading esterases, E4 and FE4, in peach-potato aphids, Myzus persicae (Sulzer), depends on both gene amplification and transcriptional control, the latter being associated with changes in DNA methylation. The structure and function of the aphid esterase genes have been studied but the determination of their copy number has proved difficult, a common problem with gene amplification. We have now used a combination of pulsed-field gel electrophoresis and quantitative competitive PCR to determine relative esterase gene copy numbers in aphid clones with different levels of insecticide resistance (R1, R2 and R3). There are approx. 4-fold increases between susceptible, R1, R2 and R3 aphids, reaching a maximum of approx. 80 times more genes in R3; this gives proportionate increases in esterase protein relative to susceptible aphids. Thus there is no overexpression of the amplified genes, in contrast with what was thought previously. For E4 genes, the loss of 5-methylcytosine is correlated with a loss of expression, greatly decreasing the amount of enzyme relative to the copy number.

Bexarotene prevents and reverses gemcitabine resistance in non-small-cell lung cancer cells by affecting gene amplification

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 2057-2057 ◽  
Author(s):  
T. W. Hermann ◽  
W. Yen ◽  
P. Tooker ◽  
S. Ng ◽  
A. Negro-Vilar ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Gene Amplification ◽  
Gene Copy Number ◽  
Dna Amplification ◽  
Fold Increase ◽  
Gene Copy ◽  
Strong Increase ◽  
Combination Regimen ◽  
Acquired Drug Resistance ◽  
Development Of Resistance

2057 Background: Bexarotene (BEX) is a retinoid X receptor selective drug approved for treatment of CTCL and has activity in various cancers. Here we evaluate its effect on acquired drug resistance towards gemcitabine (GEM) in Calu3 NSCLC models. Methods: Parental or GEM-resistant Calu3 NSCLC cells were treated with BEX and GEM as single agents or in combination. Over the treatment course the cells were evaluated for molecular changes (RNA expression and DNA amplification), which were associated with resistance status. Results: Calu3 cells treated with repeated cycles of GEM alone gradually developed resistance to this drug. However, inclusion of BEX prevented the development of resistance and cells remained chemosensitive. Microarray analysis showed a strong increase of rrm1 (ribonucleotide reductase M1) expression in the resistant cells (Calu3-GemR), a gene known to be involved in GEM resistance. In addition, the expression of genes surrounding the chromosomal location of rrm1 was increased suggesting that resistance was due to gene amplification at the 11p15 locus. This was confirmed by analysis of genomic DNA, indicating a 20-fold increase of rrm1 gene copy number. To evaluate BEX’s activity on pre-existing GEM-resistance, Calu3-GemR cells were treated with ten 10-day cycles of vehicle, GEM, BEX, or GEM plus BEX. After 4 treatment cycles the cultures treated with the combination regimen showed a loss in cell numbers, which continued until the end of the 10th cycle. This indicated a re-sensitization of the Calu3-GemR cells to treatment. The cultures treated with the single drug regimens expanded as the vehicle controls. However, analysis of RNA and DNA samples collected over the treatment course revealed that rrm1 gene amplification was gradually reduced even in the cultures treated with BEX alone, reaching parental levels at the end of the study. As expected from these data, the BEX-treated Calu3-GemR cells were again sensitive to GEM. Conclusions: The data indicate that bexarotene can re-sensitize GEM-resistant tumor cells by reversing gene amplification. This suggests that bexarotene may have general clinical utility in cancers where drug-resistance by gene amplification is a major obstacle to successful therapy. [Table: see text]

Clinical relevance of tumor cell ploidy and N-myc gene amplification in childhood neuroblastoma: a Pediatric Oncology Group study.

1991 ◽  
Vol 9 (4) ◽  
pp. 581-591 ◽  
Author(s):  
A T Look ◽  
F A Hayes ◽  
J J Shuster ◽  
E C Douglass ◽  
R P Castleberry ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Gene Amplification ◽  
Gene Copy Number ◽  
Disease Free Survival ◽  
Gene Copy ◽  
Early Treatment Failure ◽  
Free Survival ◽  
Myc Gene ◽  
Disease Free

We assessed tumor cell DNA content (ploidy) and N-myc gene copy number as predictors of long-term disease-free survival in 298 children with neuroblastoma. Diploid tumor stem lines were identified in 101 patients (34%), clonal hyperdiploid abnormalities in 194 (65%), and hypodiploid stem lines in three (1%). In children with widely disseminated tumors at diagnosis (stage D), ploidy had a highly age-dependent influence on prognosis. Among infants (less than 12 months) treated with cyclophosphamide-doxorubicin, hyperdiploidy was closely associated with long-term disease-free survival (greater than 90% of cases), while diploidy invariably predicted early treatment failure (P less than .001). Similarly, in children 12 to 24 months of age who were treated with cisplatin-teniposide and cyclophosphamide-doxorubicin, diploidy uniformly predicted early failure, whereas half of the children with hyperdiploidy achieved long-term disease-free survival (P less than .001). There was no relationship between ploidy and treatment outcome in children older than 24 months with stage D tumors who had a very low probability of long-term disease-free survival (less than 10%). N-myc gene amplification was detected in 37 (25%) of the 147 tumors tested, with the remainder showing single-copy levels of the gene. N-myc gene amplification was more frequent in diploid than in hyperdiploid tumors (23 of 57 v 14 of 87, P = .001) and predicted a high likelihood of early treatment failure. In children younger than 2 years with disseminated neuroblastoma, tumor cell ploidy and N-myc gene copy number provide complementary prognostic information that will distinguish patients who can be cured on current regimens from those who require new treatment strategies.

scholarly journals The properties of a carboxylesterase from the peach-potato aphid, Myzus persicae (Sulz.), and its role in conferring insecticide resistance

1977 ◽  
Vol 167 (3) ◽  
pp. 675-683 ◽  
Author(s):  
Alan L. Devonshire
Keyword(s):  
Insecticide Resistance ◽  
Myzus Persicae ◽  
Order Rate Constant ◽  
Preliminary Evidence ◽  
Exchange Chromatography ◽  
Potato Aphid ◽  
First Order ◽  
Different Strains ◽  
Hydrolysis Of ◽  
Peach Potato Aphid

Carboxylesterases from different strains of Myzus persicae were examined to try to understand their contribution to insecticide resistance. Preliminary evidence that they are involved comes from the good correlation between the degree of resistance and the carboxylesterase and paraoxon-degrading activity in aphid homogenates. Furthermore the carboxylesterase associated with resistance could not be separated from the insecticide-degrading enzyme by electrophoresis or ion-exchange chromatography. Homogenates of resistant aphids hydrolysed paraoxon 60 times faster than did those of susceptible aphids, yet the purified enzymes from both sources had identical catalytic-centre activities towards this substrate and also towards naphth-1-yl acetate, the latter being hydrolysed by both 2×106 times faster than paraoxon. These observations provide evidence that the enzyme from both sources is identical, and that one enzyme hydrolyses both substrates. This was confirmed by relating the rate of paraoxon hydrolysis to the rate at which paraoxon-inhibited carboxylesterase re-activated. Both had the same first-order rate constant (0.01min−1), showing clearly that the hydrolysis of both substrates is brought about by the same enzyme. Its Km for naphth-1-yl acetate was 0.131mm, and for paraoxon 75pm. The latter very small value could not be measured directly, but was calculated from substrate-competition studies coupled with measurements of re-activation of the diethyl phosphorylated enzyme. Since the purified enzymes from resistant and susceptible aphids had the same catalytic-centre activity, the 60-fold difference between strains must be caused by different amounts of the same enzyme resulting from mutations of the regulator gene(s) rather than of the structural gene.

Differential elimination of rDNA genes in bobbed mutants of Drosophila melanogaster

1986 ◽  
Vol 6 (4) ◽  
pp. 1023-1031
Author(s):  
R Terracol ◽  
N Prud'homme
Keyword(s):  
Drosophila Melanogaster ◽  
Gene Copy Number ◽  
Rdna Locus ◽  
Gene Copy ◽  
Specific Gene ◽  
Type I ◽  
28S Rrna ◽  
Wild Type ◽  
Y Chromosomes ◽  
Genes Encoding

In Drosophila melanogaster, the multiply repeated genes encoding 18S and 28S rRNA are located on the X and Y chromosomes. A large percentage of these repeats are interrupted in the 28S region by insertions of two types. We compared the restriction patterns from a subcloned wild-type Oregon R strain to those of spontaneous and ethyl methanesulfonate-induced bobbed mutants. Bobbed mutations were found to be deficiencies that modified the organization of the rDNA locus. Genes without insertions were deleted about twice as often as genes with type I insertions. Type II insertion genes were not decreased in number, except in the mutant having the most bobbed phenotype. Reversion to wild type was associated with an increase in gene copy number, affecting exclusively genes without insertions. One hypothesis which explains these results is the partial clustering of genes by type. The initial deletion could then be due either to an unequal crossover or to loss of material without exchange. Some of our findings indicated that deletion may be associated with an amplification phenomenon, the magnitude of which would be dependent on the amount of clustering of specific gene types at the locus.

scholarly journals Nosocomial Outbreak of Carbapenemase-Producing Proteus mirabilis With Two Novel Salmonella Genomic Island 1 Variants Carrying Different blaNDM–1 Gene Copies in China

2022 ◽  
Vol 12 ◽  
Author(s):  
Lang Yang ◽  
Hong He ◽  
Qichao Chen ◽  
Kaiying Wang ◽  
Yanfeng Lin ◽  
...  
Keyword(s):  
Proteus Mirabilis ◽  
Copy Number ◽  
Genomic Island ◽  
Gene Copy Number ◽  
Carbapenem Resistance ◽  
Multidrug Resistant ◽  
Gene Copy ◽  
Nosocomial Outbreak ◽  
Gene Copy Number Variation ◽  
Gene Copies

NDM-1-producing multidrug-resistant Proteus mirabilis brings formidable clinical challenges. We report a nosocomial outbreak of carbapenem-resistant P. mirabilis in China. Six P. mirabilis strains collected in the same ward showed close phylogenetic relatedness, indicating clonal expansion. Illumina and MinION sequencing revealed that three isolates harbored a novel Salmonella genomic island 1 carrying a blaNDM–1 gene (SGI1-1NDM), while three other isolates showed elevated carbapenem resistance and carried a similar SGI1 but with two blaNDM–1 gene copies (SGI1-2NDM). Four new single nucleotide mutations were present in the genomes of the two-blaNDM–1-harboring isolates, indicating later emergence of the SGI1-2NDM structure. Passage experiments indicated that both SGI variants were stably persistent in this clone without blaNDM–1 copy number changes. This study characterizes two novel blaNDM–1-harboring SGI1 variants in P. mirabilis and provides a new insight into resistance gene copy number variation in bacteria.

scholarly journals Establishing the resistance of Myzus persicae (Sulzer) by molecular methods

2008 ◽  
Vol 60 (3) ◽  
pp. 493-499 ◽  
Author(s):  
Andja Vucetic ◽  
Olivera Petrovic-Obradovic ◽  
J. Margaritopoulos ◽  
P. Skouras
Keyword(s):  
Myzus Persicae ◽  
Molecular Methods ◽  
Single Type ◽  
Heterozygous State ◽  
Potato Aphid ◽  
Serbia And Montenegro ◽  
Knock Down ◽  
Peach Potato Aphid

In two years of investigating resistance of the peach-potato aphid Myzus persicae (Sulzer) by molecular methods, several types of resistance were established in the majority of individuals from peach and tobacco in Serbia and Montenegro. Most of the tested individuals had the FE4 gene, which encodes production of FE4 esterase. The gene responsible for kdr (knock-down resistance) was found in the majority of individuals, but in the heterozygous state, while resistance based on formation of modified acetlycholinesterase (MACE) was least represented. Also, tests showed aphids from tobacco to be more sensitive to insecticide action than aphids from peach. Three tests were used in these investiga?tions, e.g., the PCR - esterase, PCR - kdr, and RFLP - PCR tests, each for a single type of resistance.

scholarly journals Effect of EPSPS Gene Copy Number and Glyphosate Selection on Fitness of Glyphosate-Resistant Bassia Scoparia in The Field

2021 ◽  
Author(s):  
Charlemagne Ajoc Lim ◽  
Prashant Jha ◽  
Vipan Kumar ◽  
Alan T. Dyer
Keyword(s):  
Sugar Beet ◽  
Seed Production ◽  
Great Plains ◽  
Copy Number ◽  
Gene Copy Number ◽  
Reproductive Traits ◽  
Gene Copy ◽  
Epsps Gene ◽  
Gene Copies

Abstract The widespread evolution of glyphosate-resistant (GR) Bassia scoparia in the U.S. Great Plains poses a serious threat to the long-term sustainability of GR sugar beet. Glyphosate resistance in B. scoparia is due to an increase in the EPSPS (5-enolpyruvyl-shikimate-3-phosphate) gene copy number. The variation in EPSPS gene copies among individuals from within a single GR B. scoparia population indicated a differential response to glyphosate selection. We tested the hypothesis of reduced GR B. scoparia fitness (reproductive traits) to increasing glyphosate rates (applied as single or sequential applications) potentially experienced within a GR sugar beet field. The variation in EPSPS gene copy number and total glyphosate rate (single or sequential applications) did not influence any of the reproductive traits of GR B. scoparia, except seed production. Sequential applications of glyphosate with a total rate of 2,214 g ae ha− 1 or higher prevented seed production in B. scoparia plants with 2–4 (low levels of resistance) and 5–6 (moderate levels of resistance) EPSPS gene copies. Timely sequential applications of glyphosate (full recommended rates) can potentially slow down the evolution of GR B. scoparia with low to moderate levels of resistance (2–6 EPSPS gene copies), but any survivors (highly-resistant individuals with ≥ 8 EPSPS gene copies) need to be mechanically removed before flowering from GR sugar beet fields. This research warrants the need to adopt ecologically based, multi-tactic strategies to reduce exposure of B. scoparia to glyphosate in GR sugar beet.

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