White Mutant Research Articles

Mapping hematopoiesis in a fully regenerative vertebrate: the axolotl

AbstractHematopoietic stem cell (HSC)-derived cells are involved in wound healing responses throughout the body. Unfortunately for mammals, wound repair typically results in scarring and nonfunctional reparation. Among vertebrates, none display such an extensive ability for adult regeneration as urodele amphibians, including 1 of the more popular models: the axolotl. However, a lack of knowledge of axolotl hematopoiesis hinders the use of this animal for the study of hematopoietic cells in scar-free wound healing and tissue regeneration. We used white and cytomegalovirus:green fluorescent protein+ transgenic white axolotl strains to map sites of hematopoiesis and develop hematopoietic cell transplant methodology. We also established a fluorescence-activated cell sorter enrichment technique for major blood lineages and colony-forming unit assays for hematopoietic progenitors. The liver and spleen are both active sites of hematopoiesis in adult axolotls and contain transplantable HSCs capable of long-term multilineage blood reconstitution. As in zebrafish, use of the white axolotl mutant allows direct visualization of homing, engraftment, and hematopoiesis in real time. Donor-derived hematopoiesis occurred for >2 years in recipients generating stable hematopoietic chimeras. Organ segregation, made possible by embryonic microsurgeries wherein halves of 2 differently colored embryos were joined, indicate that the spleen is the definitive site of adult hematopoiesis.

Identification of a hidden resistance gene in tetraploid wheat using laboratory strains of Pyricularia oryzae produced by backcrossing.

In the process (BC3F1 generation) of backcrossing an Avena isolate of Pyricularia oryzae with a Triticum isolate, color mutants with white mycelia were obtained. These white mutants lacked virulence on all (31/31) hexaploid and most (28/32) tetraploid wheat lines tested. In a BC4F1 population, white and black cultures segregated in a 1:1 ratio, suggesting that the mutant phenotype is controlled by a single gene. Furthermore, the mycelial color was perfectly linked with avirulence in the BC4F1 population; white cultures were all avirulent on common wheat (Triticum aestivum) 'Norin 4' (N4) whereas black cultures were all virulent. White cultures in the BC3F1 and BC4F1 generations were also avirulent on tetraploid wheat (T. dicoccoides) accession 'KU109' (Tat4), which was susceptible to all cultures derived from the parental wild isolates through the BC2F1 generation. A cross between Tat4 and a susceptible tetraploid (T. paleocolchicum) accession 'KU196' (Tat14) produced resistant and susceptible F2 seedlings in a 3:1 ratio against the white cultures. In the F3 generation homozygous resistant/segregating/homozygous susceptible lines segregated in a 1:2:1 ratio. These results suggest that the resistance of Tat4 to the white cultures is controlled by a single major gene. This gene, tentatively designated as RmgTd(t), is considered to be a hidden resistance gene because it was not detected with the Br58, F1, BC1F1, or BC2F1 cultures. Cytological analysis revealed that the moderate resistance controlled by RmgTd(t) was associated with a hypersensitive reaction of mesophyll cells.

Genetic modification of Methylobacterium extorquens G10 producer strain of polyhydroxybutyrate

The effect of the increased copy number of polyhydroxybutyrate (PHB) biosynthesis genes in pink-pigmented methylobacterium Methylobacterium extorquens G10 on properties of the biopolymer was studied. The activity of poly-3-hydroxybutyril-synthase (PHB-synthase) was shown to increase and the molecular weight of synthesized PHB decreases twofold (150 --> 79 kDa) after insertion of extra copies of phaC and phaCAB genes into cells of the producer strain, whereas the physicochemical properties of the plastic changed insignificantly. White mutant M. extorquens G10-W with disrupted synthesis of the carotenoid pigment (defect by the crtI gene, which codes for phytoene desaturase) was established to have the same rate of growth and level of PHB accumulation as the initial strain G10. The G10-W strain is a promising producer of PHB, with decreased expenses for purification and PHB biosynthesis.

Anatomical and genotype-specific mechanosensory responses in Drosophila melanogaster larvae

Afferent neurons commonly exhibit a somatotopic arrangement in the central nervous system that organizes spatially discrete sensory input. We are interested in how that spatial input gets integrated into motor commands. With resources for screening genes and neural circuits, and given that the cells and ion channels that transduce tactile stimuli in Drosophila larvae are remarkably well-characterized, larval mechanosensation is an ideal system for investigating how specific behaviors emerge from localized sensory input. We observed that crawling larvae are more reactive to a 20mN tactile stimulus on the head than on the tail or abdomen. Behavioral responses that were evoked by the stimuli also depended on where the stimulus was delivered. Differences in relative sensitivity were observed in different genotypes, e.g., a null white mutant and hypomorphic smn mutant are significantly more reactive to tail touches than Canton-S larvae. Responses were inhibited by silencing chemical transmission in a combination of multidendritic and chordotonal neurons, but not by inhibiting any specific subset of neurons. Extracellular recordings from segmental nerves revealed that sensory-evoked responses exhibit spike-timing dependence at the neural circuit level. Tactile stimuli reduced endogenous firing frequency and increased bursting periods when applied during periods of motor activity. The same stimulus initiated bursts of activity when applied during inactive periods. Together, these data depict the spatial and temporal complexity of mechanosensation as it applies to action selection, and provide a foundation for addressing how neural circuits in the CNS adjust their activity to afferent input.

Analysis of proteomic changes in colored mutants of Xanthophyllomyces dendrorhous (Phaffia rhodozyma)

The yeast Xanthophyllomyces dendrorhous synthesizes astaxanthin as its most prevalent xanthophyll derivative. Comparisons between the protein profiles of mutant lines of this yeast can provide insight into the carotenogenic pathway. Differently colored mutants (red, orange, pink, yellow, and white) were obtained from this yeast species, and their protein profiles were determined using two-dimensional polyacrylamide gel electrophoresis (2DE). Individual proteins differentially expressed were identified using mass spectrometry. The red mutants hyperproduced total carotenoids (mainly astaxanthin), while in white and orange mutants, mutagenesis affected the phytoene dehydrogenase activity as indicated by the accumulation of phytoene. Inactivation of astaxanthin synthase after the mutagenic treatment was evident in β-carotene accumulating mutants. Differences in the proteomic profiles of wild-type X. dendrorhous and its colored mutants were demonstrated using 2DE. Of the total number of spots detected in each gel (297-417), 128 proteins were present in all strains. The red mutant showed the greatest number of matches with respect to the wild type (305 spots), while the white and yellow mutants, which had reduced concentrations of total carotenoids, presented the highest correlation coefficient (0.6) between each other. A number of differentially expressed proteins were sequenced, indicating that tricarboxylic acid cycle and stress response proteins are closely related to the carotenogenic process.

The antimicrobial activity of pullulan film incorporated with meadowsweet flower extracts (Filipendulae ulmariae flos) on postharvest quality of apples

This study examined the antimicrobial activity of pullulan films with incorporated meadowsweet flower extracts (EMFs) and the effectiveness of these coatings in the prolonging of apple stability in various storage conditions. Pullulan was obtained from a batch culture of white mutant Aureobasidium pullulans B-1. The content of phenolic components in two extracts-water–ethanol (weEMF) and ethanol one (eEMF)-was compared using the HPLC method. Compared to eEMF, a significantly higher content was noted of phenolic acids and flavonoids in weEMF and its increased activity in inhibiting microorganism growth. Pullulan films with incorporated EMFs were obtained from the solutions to which they were added in concentrations of 3, 6, 9, 12 and 15%. These films were stable and homogenous, and to a higher degree inhibited the growth of tested bacteria than fungi. Apples of the Jonagored variety were coated with a pullulan coating containing 12% EMF. The pullulan coating with EMFs evened and unified the surface of apple skins. Pullulan coating with weEMF more effectively inhibited the growth of natural mesophilic microflora, the amount of which decreased by two logarithmic cycles at the end of storage. EMF addition to the coating significantly limited the growth of the Rhizopus arrhizus mold inoculated to the apples. EMF addition to pullulan coating did not affect color changes nor did apples loss weight during their storage. Pullulan coatings with incorporated EMFs may protect apple quality during their storage and turnover in various temperatures and time conditions.

Genetic Analysis and Gene Mapping of a Rice White Midrib Mutant wpsm

A white midrib mutant temparily designated as wpsm(white primary and secondary midrib) was screened out in the progeny of an excellent indica restorer line Jinhui 10 with seeds treated by ethyl methane sulfonate(EMS).The wpsm mutant displayed normal leaves during the seedling stage.In the late booting stage,the primary and secondary midrib of the first,second and third leaves of the wpsm mutant showed a bleached appearance,compared with those of the wild type,while mesophyll cells kept normal.This mutational character was maintained to maturity.The photosynthetic pigment of the wpsm was significantly lower than that of the wild type,so were Pn and ETR.Agronomic traits such as plant height,filled grain number per panicle,seed setting rate and 1000-grain weight were significantly decreased when compared with the wild type.Genetic analysis suggested that the mutational characters were controlled by a single recessive nuclear gene.Xinong 1A was crossed with the wpsm,892 recessive mutants from the F2 segregation population were used for gene mapping.WPSM was mapped on chromosome 6,between InDel 10 and InDel 4,with physical distance of about 56 kb.This result provides a foundation for WPSM gene cloning by map-based strategy as well as its functional analysis.

Solution structure of the strawberry allergen Fra a 1

The PR10 family protein Fra a 1E from strawberry (Fragaria x ananassa) is down-regulated in white strawberry mutants, and transient RNAi (RNA interference)-mediated silencing experiments confirmed that Fra a 1 is involved in fruit pigment synthesis. In the present study, we determined the solution structure of Fra a 1E. The protein fold is identical with that of other members of the PR10 protein family and consists of a seven-stranded antiparallel β-sheet, two short V-shaped α-helices and a long C-terminal α-helix that encompass a hydrophobic pocket. Whereas Fra a 1E contains the glycine-rich loop that is highly conserved throughout the protein family, the volume of the hydrophobic pocket and the size of its entrance are much larger than expected. The three-dimensional structure may shed some light on its physiological function and may help to further understand the role of PR10 proteins in plants.

Characterization of a heavy-ion induced white flower mutant of allotetraploid Nicotiana tabacum

KEY MESSAGE : We characterized a white flower mutant of allotetraploid N. tabacum as a DFR-deficient mutant; one copy of DFR has a cultivar-specific frameshift, while the other was deleted by heavy-ion irradiation. In most plants, white-flowered mutants have some kind of deficiency or defect in their anthocyanin biosynthetic pathway. Nicotiana tabacum normally has pink petals, in which cyanidin is the main colored anthocyanidin. When a relevant gene in the cyanidin biosynthetic pathway is mutated, the petals show a white color. Previously, we generated white-flowered mutants of N. tabacum by heavy-ion irradiation, which is accepted as an effective mutagen. In this study, we determined which gene was responsible for the white-flowered phenotype of one of these mutants, cv. Xanthi white flower 1 (xwf1). Southern blot analysis using a DNA fragment of the dihydroflavonol 4-reductase (DFR) gene as a probe showed that the xwf1 mutant lacked signals that were present in wild-type genomic DNAs. Sequence analysis demonstrated that one copy of the DFR gene (NtDFR2) was absent from the genome of the xwf1 mutant. The other copy of the DFR gene (NtDFR1) contained a single-base deletion resulting in a frameshift mutation, which is a spontaneous mutation in cv. Xanthi. Introduction of NtDFR2 cDNA into the petal limbs of xwf1 by particle bombardment resulted in production of the pink-colored cells, whereas introduction of NtDFR1 cDNA did not. These results indicate that xwf1 is a DFR-deficient mutant. One copy of NtDFR1 harbors a spontaneous frameshift mutation, while the other copy of NtDFR2 was deleted by heavy-ion beam irradiation.

Intragenic Enhancers and Suppressors of Phytoene Desaturase Mutations in Chlamydomonas reinhardtii

Photosynthetic organisms synthesize carotenoids for harvesting light energy, photoprotection, and maintaining the structure and function of photosynthetic membranes. A light-sensitive, phytoene-accumulating mutant, pds1-1, was isolated in Chlamydomonas reinhardtii and found to be genetically linked to the phytoene desaturase (PDS) gene. PDS catalyzes the second step in carotenoid biosynthesis—the conversion of phytoene to ζ-carotene. Decreased accumulation of downstream colored carotenoids suggested that the pds1-1 mutant is leaky for PDS activity. A screen for enhancers of the pds1-1 mutation yielded the pds1-2 allele, which completely lacks PDS activity. A second independent null mutant (pds1-3) was identified using DNA insertional mutagenesis. Both null mutants accumulate only phytoene and no other carotenoids. All three phytoene-accumulating mutants exhibited slower growth rates and reduced plating efficiency compared to wild-type cells and white phytoene synthase mutants. Insight into amino acid residues important for PDS activity was obtained through the characterization of intragenic suppressors of pds1-2. The suppressor mutants fell into three classes: revertants of the pds1-1 point mutation, mutations that changed PDS amino acid residue Pro64 to Phe, and mutations that converted PDS residue Lys90 to Met. Characterization of pds1-2 intragenic suppressors coupled with computational structure prediction of PDS suggest that amino acids at positions 90 and 143 are in close contact in the active PDS enzyme and have important roles in its structural stability and/or activity.

Flower color mutant and its possible significance in Commelina benghalensis L. (Commelinaceae)

Commelina benghalensis L. bears blue to blue-violet flowers. A few plants of a white flowering mutant found growing naturally at three different places in Jammu were transplanted in the Botanical Garden of University of Jammu. The two color forms were subjected to critical morphological and reproductive analyses. Apart from significant differences in only a few traits, the two flower types were largely similar in most of the morphological features and were true breeding. F1 hybrids obtained had blue flowers and were adequately fertile. F2 hybrids segregating in the ratio of 3(blue):1(white) were equally fertile. Results of χ 2 test indicate monohybrid inheritance of flower color with blue completely dominant over white. Besides this, the possible role of flower color variation in the species is also discussed.

Use of β-galactosidase (lacZ) gene α-complementation as a novel approach for assessment of titanium oxide nanoparticles induced mutagenesis

The mutagenic potential of titanium dioxide nanoparticles (TiO2-NPs) of an average size 30.6nm was investigated using β-galactosidase (lacZ) gene complementation in plasmid pUC19/lacZ−Escherichia coli DH5α system. Plasmid pUC19 was treated with varying concentrations of TiO2-NPs and allowed to transfect the CaCl2-induced competent DH5α cells. The data revealed loss in transformation efficiency of TiO2-NPs treated plasmids as compared to untreated plasmid DNA in DH5α host cells. Induction of multiple mutations in α-fragment of lacZ gene caused synthesis of non-functional β-galactosidase enzyme, which resulted in a significant number of white (mutant) colonies of transformed E. coli cells. Screening of mutant transformants based on blue:white colony assay and DNA sequence analysis of lacZ gene fragment clearly demonstrated TiO2-NPs induced mutagenesis. Multiple alignment of selectable marker lacZ gene sequences from randomly selected mutants and control cells provided a gene specific map of TiO2-NPs induced mutations. Mutational analysis suggested that all nucleotide changes were point mutations, predominantly transversions (TVs) and transitions (TSs). A total of 32 TVs and 6 TSs mutations were mapped within 296 nucleotides (nt) long partial sequence of lacZ gene. The region between 102 and 147nt within lacZ gene sequence was found to be most susceptible to mutations with nine detectable point mutations (8 TVs and 1 TSs). Guanine base was determined to be more prone to TiO2-NPs induced mutations. This study suggested the pUC19/E. coli DH5αlacZ gene α-complementation system, as a novel genetic approach for determining the mutagenic potential, and specificity of manufactured NPs and nanomaterials.

White Mutants of Chloroperoxidase-Secreting Caldariomyces fumago as Superior Production Strains, Revealing an Interaction between Pigmentation and Enzyme Secretion

By mutant-colony screening of Caldariomyces fumago, several white mutants were isolated that are superior strains for the production of the valuable enzyme chloroperoxidase (CPO). Their culture supernatant lacks the contaminating dark pigment, which simplifies downstream processing. Furthermore, the CPO content increased significantly faster than the wild-type rate, which uncovers possible interactions between pigmentation and enzyme secretion.

Backbone and side chain 1H, 13C, and 15N Chemical Shift Assignments for Hen Egg White Lysozyme mutant WT-ALA

Backbone and side chain 1H, 13C, and 15N Chemical Shift Assignments for Hen Egg White Lysozyme mutant WT-ALA

Differentially expressed genes in white egg 2 mutant of silkworm, Bombyx mori, at early embryo development stages

White egg 2 is one of white egg mutants in silkworm, whose molecular mechanism remains unknown so far. In order to obtain an overall view on gene expression profiles at early embryo development stages, the white egg 2 near-isogenic line was constructed and the whole-genome of silkworm microarray system containing 21375 predicted genes from the silkworm whole genome sequence was employed to investigate gene expression profiles at 0, 24 and 48 h post oviposition between white egg 2 mutant and normal black egg strain. At 24 h post oviposition, 49 genes exhibited at least 2.0 fold differences at expression level, including 24 up-regulated genes and 25 down-regulated genes while at 48 h post oviposition, 52 genes, including 23 up-regulated genes and 29 down-regulated genes were expressed differentially over 2.0 change fold. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that nine differentially expression genes were involved in nine significant (p<0.05) pathways at 24 h post oviposition and 24 significant pathways at 48 h post oviposition, respectively. These pathways were related to amino acid metabolism, sugar metabolism, and series of major physiological metabolism. Our results hopefully shed light on the further study of molecular mechanism of white egg 2 mutant. Key words : Bombyx mori , white egg 2 mutant, microarray, embryo, differentially expressed gene.

Defining the Axolotl Hematopoietic Stem Cell

Abstract 1295Hematopoietic stem cells (HSCs) are the most therapeutically usable stem cells to date. HSC derived cells are involved in wound healing responses throughout the body. The goal of our study is to discovery methods to promote improved regenerative responses from mammalian HSC. The champion of vertebrate regeneration is the axolotl. It can regrow entire limbs, major portions of most internal organs – including the brain and spinal cord. Recent advances in the production of transgenic axolotl, complete mapping of the axolotl transcriptome and production of gene expression arrays finally allow molecular mapping of regeneration pathways. In order to define the role of HSC in regeneration it was first necessary to define and characterize the axolotl HSC. In the current study we develop classic assays such as hematopoietic cell transplantations (HCT) and colony forming cell (CFC) assays to map axolotl hematopoiesis. In addition we take advantage of embryo manipulations possible with the axolotl to generate hematopoietic chimeras (GFP+ vs. white) as non-injury models for regeneration.Axolotl have a naturally occurring white variant that are nearly as transparent as zebrafish. We used a normal and CMV:GFP+ transgenic white strains to map sites of hematopoiesis and to develop HSC transplant methodology. We also establish protocols for Colony Forming Unit (CFU) assays for hematopoietic progenitors. Liver, spleen, kidney, marrow, and thymus were tested as potential HSC sources. Both the liver and spleen contained transplantable HSC capable of radioprotection and multilineage reconstitution of lethally irradiated (1000 rads) adult axolotls or non-conditioned embryos. As in zebrafish, use of the white axolotl mutant allows direct visualization of engraftment, homing, and hematopoiesis in real time. Fluorescent donor cells were seen homing to the liver and spleen regardless of tissue origin, albeit with better overall engraftment from spleen cells. Hematopoiesis occurred for more than 6 months in embryo transplantations. FACS analysis of these two organs showed that the liver contains relatively even numbers of myeloid and lymphoid populations whereas the spleen has a bias toward lymphoid and erythroid populations. Additionally, early stage white and GFP embryos were transected and the posterior and anterior halves of the two different animals were joined. GFP+ blood was present in animals that contained the liver or spleen within the GFP portion of the body further indicating that hematopoiesis occurs in these organs. These embryo chimeras also provide stable long-term GFP+ blood chimeras without irradiation injury. CFU assays demonstrated that the liver contains predominantly myeloid progenitors. In contrast, the spleen has mostly erythroid and megakaryocytic progenitors but also some myeloid progenitors.Here we have shown that the axolotl HSC resides in the liver and spleen and begun the basic mapping of hematopoiesis. Moreover, we have established the essential hematopoietic assays for the axolotl. The ability to generate both irradiation and non-injury GFP+ blood chimeras provide powerful tools to determine the role of HSC/progeny in axolotl regeneration. Disclosures:No relevant conflicts of interest to declare.

Functional Analysis of the Phycomyces carRA Gene Encoding the Enzymes Phytoene Synthase and Lycopene Cyclase

Phycomyces carRA gene encodes a protein with two domains. Domain R is characterized by red carR mutants that accumulate lycopene. Domain A is characterized by white carA mutants that do not accumulate significant amounts of carotenoids. The carRA-encoded protein was identified as the lycopene cyclase and phytoene synthase enzyme by sequence homology with other proteins. However, no direct data showing the function of this protein have been reported so far. Different Mucor circinelloides mutants altered at the phytoene synthase, the lycopene cyclase or both activities were transformed with the Phycomyces carRA gene. Fully transcribed carRA mRNA molecules were detected by Northern assays in the transformants and the correct processing of the carRA messenger was verified by RT-PCR. These results showed that Phycomyces carRA gene was correctly expressed in Mucor. Carotenoids analysis in these transformants showed the presence of ß-carotene, absent in the untransformed strains, providing functional evidence that the Phycomyces carRA gene complements the M. circinelloides mutations. Co-transformation of the carRA cDNA in E. coli with different combinations of the carotenoid structural genes from Erwinia uredovora was also performed. Newly formed carotenoids were accumulated showing that the Phycomyces CarRA protein does contain lycopene cyclase and phytoene synthase activities. The heterologous expression of the carRA gene and the functional complementation of the mentioned activities are not very efficient in E. coli. However, the simultaneous presence of both carRA and carB gene products from Phycomyces increases the efficiency of these enzymes, presumably due to an interaction mechanism.

The Kynurenine Pathway Modulates Neurodegeneration in a Drosophila Model of Huntington's Disease

Neuroactive metabolites of the kynurenine pathway (KP) of tryptophan degradation have been implicated in the pathophysiology of neurodegenerative disorders, including Huntington's disease (HD) [1]. A central hallmark of HD is neurodegeneration caused by a polyglutamine expansion in the huntingtin (htt) protein [2]. Here we exploit a transgenic Drosophila melanogaster model of HD to interrogate the therapeutic potential of KP manipulation. We observe that genetic and pharmacological inhibition of kynurenine 3-monooxygenase (KMO) increases levels of the neuroprotective metabolite kynurenic acid (KYNA) relative to the neurotoxic metabolite 3-hydroxykynurenine (3-HK) and ameliorates neurodegeneration. We also find that genetic inhibition of tryptophan 2,3-dioxygenase (TDO), the first and rate-limiting step in the pathway, leads to a similar neuroprotective shift toward KYNA synthesis. Importantly, we demonstrate that the feeding of KYNA and 3-HK to HD model flies directly modulates neurodegeneration, underscoring the causative nature of these metabolites. This study provides the first genetic evidence that inhibition of KMO and TDO activity protects against neurodegenerative disease in an animal model, indicating that strategies targetedattwo key points within the KP may have therapeutic relevance in HD, and possibly other neurodegenerative disorders.

White Mutants of Straight Edge: The Avant-Garde of Abstinence

The Journal of Popular CultureVolume 44, Issue 3 p. 633-646 White Mutants of Straight Edge: The Avant-Garde of Abstinence GABRIEL SMITH, GABRIEL SMITH Indiana University of PennsylvaniaSearch for more papers by this author GABRIEL SMITH, GABRIEL SMITH Indiana University of PennsylvaniaSearch for more papers by this author First published: 02 June 2011 https://doi.org/10.1111/j.1540-5931.2011.00852.xCitations: 3Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume44, Issue3June 2011Pages 633-646 RelatedInformation

Isolation of &lt;em&gt;Drosophila melanogaster&lt;/em&gt; Testes

The testes of Drosophila melanogaster provide an important model for the study of stem cell maintenance and differentiation, meiosis, and soma-germline interactions. Testes are typically isolated from adult males 0-3 days after eclosion from the pupal case. The testes of wild-type flies are easily distinguished from other tissues because they are yellow, but the testes of white mutant flies, a common genetic background for laboratory experiments are similar in both shape and color to the fly gut. Performing dissection on a glass microscope slide with a black background makes identifying the testes considerably easier. Testes are removed from the flies using dissecting needles. Compared to protocols that use forceps for testes dissection, our method is far quicker, allowing a well-practiced individual to dissect testes from 200-300 wild-type flies per hour, yielding 400-600 testes. Testes from white flies or from mutants that reduce testes size are harder to dissect and typically yield 200-400 testes per hour.