Sequence analysis of lacZ~- mutations induced by ion beam irradiation in double-stranded M13mp18DNA

While M13mpl8 double-stranded DNA was irradiated with ion beam, and transfected into E. coli JM103, a decrease of transfecting activity was discovered. The lacZ-mutation frequency at 20% survival could reach (3.6-16.8) × 104, about 2.3-10 times that of unirradiated M13DNA. Altogether, 27 lacZ~ mutants were select-ed, 10 of which were used for sequencing. 7 of the sequenced mutants show base changes in 250-bp region examined (the remaining 3 mutants probably have base changes outside the regions sequenced). 5 of the base-changed mutants contain more than one mutational base sites (some of them even have 5-6 mutational base sites in 250-bp region ex-amined) ; this dense distribution of base changes in polysites has seldom been seen in X-rays, γ-rays or UV induced DNA mutations. Our experiments also showed that the types of base changes include transitions( 50 % ), transversions (45% ) and deletion (5% ); no addition or duplication was observed. The transitions were mainly C→T and A→G; the transversion     

Cloning, sequencing and function ofsanA, a gene involved in nikkomycin biosynthesis ofStreptomyces ansochromogenes

Several genetically stable mutants blocked in nikkomycin biosynthesis were obtained after the slightly germinated spores of Streptomyces ansochromogenes, a nikkomycin producer, were treated with ultra violet radiation. One of the mutants is the same in morpholotical differentiation as the wild type strain and is designated as NBB19. A DMA library was constructed using plasmid plJ702 as cloning vector, NBB19 as cloning recipient. A 6 kb DNA fragment which can genetically complement NBB19 was cloned when screening the library for antifungal activity. Sequence analysis showed that the 3 kb Bgl II-Sal I fragment contains one complete ORF (ORF1) and one partial ORF (ORF2). ORF1 is designated as sanA. sanA is 1 365 bp, encoding a protein consisting of 454 amino acid residues. Database searching indicated that sanA is homologous to the hypothetical methyltransferase in Pyrococcus horikoshli with 25% identities and 41% positives. Disruptant of sanA lost the ability to synthesize nikkomycin. It indicated that sa     

cvhA gene of Streptomyces hygroscopicus 10-22 encodes a negative regulator for mycelia development

A five-gene cluster cvhABCDE was identified from Streptomyces hygroscopicus 10-22.As thefirst gene of this cluster,cvhA encoded a putative sensor histidine kinase with a predicted sensor domainconsisting of two trans-membrane segments at the N-terminus and a conserved HATPase_c domain at the C-terminus.The C-terminus polypeptide of CvhA expressed in Escherichia coli was purified and shown to beautophosphorylated with [γ-~(32)P]ATP in vitro.The phosphoryl group was acid-labile and basic-stable,whichsupported histidine as the phosphorylation residue.No obvious difference of mycelia development was ob-served between the null mutant of cvhA generated by targeted gene replacement and the wild-type parentalstrain 10-22 grown on solid soya flour medium with 2%-8% glucose or sucrose,but the cvhA mutant couldform much more abundant aerial mycelia and spores than the wild-type strain on solid soya flour mediumsupplemented with 6%-8% mannitol,6%-8% sorbitol,4%-6% mannose,or 4%-6% fructose.This pheno-type was complemented by the cloned wild-type cvhA gene,and no difference was observed for growthcurves of the cvhA mutant and the wild strain in liquid minimal medium with the tested sugars at a concen-tration of 4%,6% and 8%.We thus propose that CvhA is likely a sensor histidine kinase and negativelyregulates the morphological differentiation in a sugar-dependent manner in S.hygroscopicus 10-22.     

Influence of growing location and cultivar on Rhyzopertha dominica （Coleoptera： Bostrichidae） and Sitophilus oryzae （Coleoptera： Curculionidae） infestation of rough rice

Long-grain rice cultivars Cocodrie, Wells, and XP 723 grown in three locations （Hazen, MO; Essex and Newport, AR, USA）, and medium-grain rice cultivars Bengal and XP 713 grown in two locations （Jonesboro and Lodge Corner, AR, USA）, were harvested and assayed for susceptibility to Rhyzopertha dominica （F.） （Coleoptera： Bostrichidae）, the lesser grain borer, and Sitophilus oryzae （L.） （Coleoptera： Curculionidae）, the rice weevil, on rice held at 27℃, 57% and 75% relative humidity （RH）. Separate samples from the same harvest lots were also analyzed for the physical characteristics of brown rice yield, percentage whole kernels and kernel thickness. Progeny production and feeding damage of R. dominica were significantly different among long-grain cultivars within two of the three locations （P 〈 0.05）, but not for location or RH （P ≥ 0.05）, while progeny production of S. oryzae was different among cultivars, location, and RH （P 〈 0.05）. On medium-grain rice, both cultivar and location were significant for progeny production of R. dominica, but not RH, while cultivar and RH were significant for progeny production of S. oryzae, but not location. On both rice types, feeding damage of R. dominica followed the same trends and was always strongly positively correlated with progeny production （P 〈 0.05）, but for S. oryzae there were several instances in which progeny production was not correlated with feeding damage （P ≥ 0.05）. Physical characteristics of both rice types were statistically significant （P 〈 0.01） but actual numerical differences were extremely small, and were generally not correlated with progeny production of either species. Results indicate that the location in which a particular rice cultivar is grown, along with its characteristics, could affect susceptibility of the rice to R. dominica and S. oryzae.     

Effect of different immunosuppressive drugs on calcineurin and its mutants

Several mutants in Loop7 region and near Loop7 region of calcineurin A (CN A) subunit have been constructed and purified using site-directed mutagenesis. Their phosphatase activity and the corresponding solution conformation were examined. Their phosphatase activities between wild-type CN and mutants were compared to identify the interaction of different immuno-suppressive drugs with CN. The results showed that the phosphatase activities of the mutants at Loop7 were much higher than the one of wild-type CN. Furthermore, circular dichroism spectra of the mutants revealed that their solution conformations gave rise in changes in native structure of the protein. Cyclophilin-CyclosporinA (CyP-CsA) significantly inhibited the phosphatase activity of wild-type CN, and had no effects on the phosphatase activity of mutants in Loop7 region, which indicates that the site-directed mutagenesis at Loop7 region made a significant change in the interaction between CyP-CsA and CN. Examination of the activities of these     

Two distinct conjugal transfer systems on Streptomyces plasmid pZL1

The mechanism of conjugal transfer of plasmids in Gram-negative and unicellular Gram-positive bacteria is commonly via a type IV secretion system （T4SS） [1]. The genes encoding the T4S proteins are usually arranged in a single operon or a few operons. In Gram-negative Agrobacterium tumefaciens, the T4SS is encoded by the virB and virD operons （11 and 5 genes, respectively） [2]. Streptomyces are multicellular mycelial Gram-positive bac- teria that form unicellular spores. There are fundamental dif- ferences in the mechanisms of conjugal transfer between Streptomyces plasmids and those of Gram-negative and uni- cellular Gram-positive bacteria. Conjugal transfer of Streptomyces plasmids requires a tra gene encoding an FtsK/SpolIIE-family DNA translocator and a few adjacent genes [3]. No bacterial T4SS has previously been found on Streptomyces conjugative plasmids. We reported here the co- existence of both a T4SS-like and an FtsK/SpolIIE-family DNA translocator on a 128-kb Streptomyces plasmid, pZL 1.     

Antioxidation of Anthocyanins in Photosynthesis Under High Temperature Stress

Chlorophyll fluorescence and antioxidative capability in detached leaves of the wild type Arabidopsis thaliana L.ecotypeLandsberg erecta(Ler)and three mutants deficient in anthocyanins biosynthesis(tt3,tt4,and tt3tt4)were investigatedduring treatment with temperatures ranging 25-45℃.In comparison with the wild type,chlorophyll fluorescence parametersFv/Fm,Φ_(PSII),electron transport rate(ETR),Fv/Fo and qP in three anthocyanin-deficient mutants showed a more rapidlydecreasing rate when the temperature was over 35℃.Non-photochemical quenching(NPQ)in these mutants was almostcompletely lost at 44℃,whereas the content of heat stable protein dropped and the rate of the membrane leakage increased.Fo-temperature curves were obtained by monitoring Fo levels with gradually elevated temperatures from 22℃ to 72℃ at0.5℃/min.The inflexion temperatures of Fo were 45.8℃ in Ler,45.1℃ in tt3,44.1℃ in tt4 and 42.3℃ in tt3tt4,respectively.The temperatures of maximal Fo in three mutants were 1.9-3.8℃ lower than the wild type plants.Meanwhile,three mutantshad lower activities of superoxide dismutase(SOD)and ascorbate peroxidase(APX)and an inferior scavenging capabilityto DPPH(1.1-diphenyl-2-picrylhy.drazyl)radical under heat stress,and in particular tt3tt4 had the lowest antioxidativepotential.The results of the diaminobenzidine-H_2O_2 histochemical staining showed that H_2O_2 was accumulated in theleaf vein and mesophyll cells of mutants under treatment at 40℃,and it was significantly presented in leaf cells of tt3tt4.The sensitivity of Arabidopsis anthocyanins-deficient mutants to high temperatures has revealed that anthocyanins innormal plants might provide protection from high temperature injury,by enhancing its antioxidative capability under hightemperature stress.     

Antioxidation of Anthocyanins in Photosynthesis Under High Temperature Stress

Chlorophyll fluorescence and antioxidative capability in detached leaves of the wild type Arabidopsis thaliana L. ecotype Landsberg erecta （Ler） and three mutants deficient in anthocyanins biosynthesis （tt3, tt4, and tt3tt4） were investigated during treatment with temperatures ranging 25-45 ℃. In comparison with the wild type, chlorophyll fluorescence parameters Fv/Fm, φps,, electron transport rate （ETR）, Fv/Fo and qP in three anthocyanin-deficient mutants showed a more rapidly decreasing rate when the temperature was over 35 ℃. Non-photochemical quenching （NPQ） in these mutants was almost completely lost at 44 ℃, whereas the content of heat stable protein dropped and the rate of the membrane leakage increased. Fo-temperature curves were obtained by monitoring Fo levels with gradually elevated temperatures from 22 ℃ to 72 ℃ at 0.5 ℃/min. The inflexion temperatures of Fo were 45.8 ℃ in Ler, 45.1℃ in tt3, 44.1℃ in tt4 and 42.3 ℃ in tt3tt4, respectively. The temperatures of maximal Fo in three mutants were 1.9-3.8℃ lower than the wild type plants. Meanwhile, three mutants had lower activities of superoxide dismutase （SOD） and ascorbate peroxidase （APX） and an inferior scavenging capability to DPPH （1.1-diphenyl-2-picrylhy.drazyl） radical under heat stress, and in particular tt3tt4 had the lowest antioxidative potential. The results of the diaminobenzidine-H2O2 histochemical staining showed that H2O2 was accumulated in the leaf vein and mesophyll cells of mutants under treatment at 40 ℃, and it was significantly presented in leaf cells of tt3tt4. The sensitivity of Arabidopsis anthocyanins-deficient mutants to high temperatures has revealed that anthocyanins in normal plants might provide protection from high temperature injury, by enhancing its antioxidative capability under high temperature stress.     

The response of electron transport mediated by active NADPH dehydrogenase complexes to heat stress in the cyanobacterium Synechocystis 6803

The electron-transport machinery in photosynthetic membranes is known to be very sensitive to heat. In this study, the rate of electron transport (ETR) driven by photosystem I (PSI) and photosystem II (PSII) during heat stress in the wild-type Synechocystis sp. strain PCC 6803 (WT) and its ndh gene inactiva-tion mutants △ndhB (M55) and △ndhD1/ndhD2 (D1/D2) was simultaneously assessed by using the novel Dual-PAM-100 measuring system. The rate of electron transport driven by the photosystems (ETRPSs) in the WT, M55, and D1/D2 cells incubated at 30℃ and at 55℃ for 10 min was compared. Incubation at 55 ℃ for 10 min significantly inhibited PSII-driven ETR (ETRPSII) in the WT, M55 and D1/D2 cells, and the ex-tent of inhibition in both the M55 and D1/D2 cells was greater than that in the WT cells. Further, PSI-driven ETR (ETRPSI) was stimulated in both the WT and D1/D2 cells, and this rate was increased to a greater extent in the D1/D2 than in the WT cells. However, ETRPSI was considerably inhibited in the M55 cells. Analysis of the effect of heat stress on ETRPSs with regard to the alterations in the 2 active NDH-1 complexes in the WT, M55, and D1/D2 cells indicated that the active NDH-1 supercomplex and medi-umcomplex are essential for alleviating the heat-induced inhibition of ETRPSII and for accelerating the heat-induced stimulation of ETRPSI, respectively. Further, it is believed that these effects are most likely brought about by the electron transport mediated by each of these 2 active NDH-1 complexes.     

The two major spore DNA repair pathways, nucleotide excision repair and spore photoproduct lyase, are sufficient for the resistance of Bacillus subtilis spores to artificial UV-C and UV-B but not to solar radiation.

Bacterial endospores are 1 to 2 orders of magnitude more resistant to 254-nm UV (UV-C) radiation than are exponentially growing cells of the same strain. This high UV resistance is due to two related phenomena: (i) DNA of dormant spores irradiated with 254-nm UV accumulates mainly a unique thymine dimer called the spore photoproduct (SP), and (ii) SP is corrected during spore germination by two major DNA repair pathways, nucleotide excision repair (NER) and an SP-specific enzyme called SP lyase. To date, it has been assumed that these two factors also account for resistance of bacterial spores to solar UV in the environment, despite the fact that sunlight at the Earth's surface consists of UV-B, UV-A, visible, and infrared wavelengths of approximately 290 nm and longer. To test this assumption, isogenic strains of Bacillus subtilis lacking either the NER or SP lyase DNA repair pathway were assayed for their relative resistance to radiation at a number of UV wavelengths, including UV-C (254 nm), UV-B (290 to 320 nm), full-spectrum sunlight, and sunlight from which the UV-B portion had been removed. For purposes of direct comparison, spore UV resistance levels were determined with respect to a calibrated biological dosimeter consisting of a mixture of wild-type spores and spores lacking both DNA repair systems. It was observed that the relative contributions of the two pathways to spore UV resistance change depending on the UV wavelengths used in a manner suggesting that spores irradiated with light at environmentally relevant UV wavelengths may accumulate significant amounts of one or more DNA photoproducts in addition to SP. Furthermore, it was noted that upon exposure to increasing wavelengths, wild-type spores decreased in their UV resistance from 33-fold (UV-C) to 12-fold (UV-B plus UV-A sunlight) to 6-fold (UV-A sunlight alone) more resistant than mutants lacking both DNA repair systems, suggesting that at increasing solar UV wavelengths, spores are inactivated either by DNA damage not reparable by the NER or SP lyase system, damage caused to photosensitive molecules other than DNA, or both.     

Escherichia coli Mutants Tolerant to Beta-Lactam Antibiotics

Two types of Escherichia coli mutants tolerant to beta-lactam antibiotics were isolated. One is E. coli chi2452, which showed a tolerant response against beta-lactam antibiotics when grown at 42 degrees C, and the others are the mutants C-80 and C-254, selected from mutagenized E. coli chi1776 by cycles of exposure to ampicillin, cephaloridine, and starvation of the nutritionally required diaminopimelic acid. Beta-lactam antibiotics caused rapid loss of viability and lysis in cultures of chi1776 or in chi2452 grown at 32 degrees C. In contrast, the same antibiotics caused only a reversible inhibition of growth in mutants C-80 and C-254 or in cultures of chi2452 grown at 42 degrees C. Beta-lactam antibiotics that show high affinity for penicillin-binding proteins 2 or 3 (mecillinam and cephalexin, respectively) induced similar morphological effects (ovoid cell formation and filament formation) in both parent and mutant strains. In contrast, beta-lactam antibiotics which have a high affinity for penicillin-binding protein 1 (e.g., cephaloridine or cefoxitin), which cause rapid lysis in the parental strains, caused cell elongation in the tolerant bacteria. In contrast to the parental cells, autolytic cell wall degradation was not triggered by beta-lactam treatment of chi2452 cells grown at 42 degrees C or in mutants C-80 and C-254. The total autolytic activity of mutants C-80 and C-254 was less than 30% that of the parent strain. However, virtually identical autolytic activities were found in cells of chi2452 grown either at 42 or 32 degrees C. Possible mechanisms for the penicillin tolerance of E. coli are considered on the basis of these findings.     

Use of Streptomyces sp. 26-115 protoplasts inactivated by heating in fusion experiments

When heated at 55 degrees C for 30 or 60 minutes protoplasts of auxotrophic mutants of Streptomyces sp. 26-115 producer of actinomycin C (active and inactive variants) lost their capacity for regeneration. The protoplasts heated at at 55 degrees C for 30 minutes and not for 60 minutes maintained some ability to yield recombinants on fusion under the effect of PEG 6000. Unlike the parent active strain, the colonies formed by the spores of the prototrophs yielding on fusion of the intact protoplasts showed wide ranges of antibiotic activity against M. flavus while a significant part of the colonies was inactive. The use of the inactive variant protoplasts heated at 55 degrees C for 30 minutes in the fusion procedure increased the proportion of the inactive variants.     

Transitory germinative excision repair in Bacillus subtilis.

Bacillus subtilis strains UVSSP-42-1 (hcr42 ssp1) and UVSSP-1-1 (hcr1 ssp1) are ultraviolet (UV) radiation sensitive both as dormant spores and as vegetative cells. These strains are unable to excise cyclobutane-type dimers from the deoxyribonucleic acid (DNA) of irradiated vegetative cells and fail to remove spore photoproduct from the DNA of irradiated spores either by excision (controlled by gene hcr) or by spore repair (controlled by gene ssp1). When irradiated soon after spore germination, these strains excise dimers, but not spore photoproduct, from their DNA. This process, termed germinative excision repair, functions only transiently in the germination phase and is responsible for the high UV resistance of germinated spores and for their temporary capacity to host cell reactivate irradiated phages infecting them. The recA1 mutation confers higher UV sensitivity to the germinated spores, but does not interfere with dimer removal by germinative excision repair.     

阿维链霉菌的诱变育种

以阿维链霉菌(Streptomyces avermitilis)76-12为出发菌株,采用亚硝基胍、吖啶橙、紫外线和氯化锂分别对其孢子和原生质体进行诱变,经抗代谢物理性筛选,获得一系列高产突变株,其中N-1-2高产突变株的发酵单位是出发菌株的2.47倍。实验中同时获得了只产阿维菌素a组分的突变株G-32、Bla组分含量高的Ave8菌株和产蓝绿色孢子的突变株UA-G等。     

链霉菌6803菌株对植物的化感作用

链霉菌属是绝大多数已知抗生素和一些重要活性物质的产生菌,它对高等植物是否具有化感作用尚缺乏研究. 从土壤中分离获得7个放线菌菌株,研究其对植物幼苗生长的抑制作用.结果表明： 链霉菌6803菌株在固体和液体发酵培养时均强烈抑制油菜根和稗草根的生长, 其液体发酵液的5倍稀释液对油菜和稗草苗生长的抑制分别达到60.7%和61.3%. 常规形态与生理生化试验表明,该菌株属于链霉菌属金色类群; 16S rRNA基因序列分析表明,该菌株为沙场链霉菌,基因序列相似性达到99.28%. 此菌株经过80～100 s紫外线照射筛选出的正突变菌株UV8024和UV100-2的10倍稀释发酵液对油菜幼苗生长的抑制作用比原始菌株分别提高了37.5%和38.1%, 经过1%硫酸二乙酯诱变50 min筛选出的正突变菌株D507的10倍稀释发酵液对油菜幼苗生长的抑制作用比原始菌株提高了29.8%. 链霉菌6803菌株对高等植物具有化感作用, 并可通过诱变育种提高其化感潜力.     

Direct Enzymatic Repair of Deoxyribonucleic Acid Single-Strand Breaks in Dormant Spores

With the alkaline sucrose gradient centrifugation method, it was found that dormant spores of Clostridium botulinum subjected to 300 krads of gamma radiation showed a distinct decrease in deoxyribonucleic acid (DNA) fragment size, indicating induction of single-strand breaks (SSB). A two- to threefold difference in radiation resistance of spores of two strains of C. botulinum, 33A (37% survival dose [D(37)] = 110 krads) and 51B (D(37) = 47 krads), was accompanied by relatively larger DNA fragments (molecular weight 7.9 x 10(7)) obtained during extraction from the radiation-resistant strain 33A and smaller DNA fragments (molecular weight 1.8 x 10(7)) obtained under identical conditions from radiation-sensitive strain 51B. The apparent number of DNA SSB produced by 300 krads in strains 33A and 51B was 0.37 and 3.50, respectively, per 10(8) daltons of DNA. Addition of 0.02 M ethylenediaminetetraacetic acid (EDTA) to spore suspensions during irradiation doubled the apparent number of SSB in strain 33A but had no effect on strain 51B. In vivo, 0.02 M EDTA present during irradiation to 100 to 300 krads decreased survival of spores of 33A by about 30% but had little or no effect on 51B. Survival of 33A was also reduced by about 45% when the spores were irradiated while frozen in dry ice (-75 C) and, after irradiation, immediately exposed to 0.03 M EDTA for 1 h to inhibit repair in the dormant spores. These results suggest that the highly radiation-resistant strain 33A may be able to accomplish repair of SSB during irradiation or after irradiation under nonphysiological conditions, i.e., in the dormant state. This repair can be inhibited by EDTA. Sedimentation patterns show that DNA from spores of both strains 33A and 51B did not show any postirradiation repair during the first 6 h of germination, as opposed to Bacillus subtilis spores, which exhibit repair immediately after germination. These observations suggest the existence of direct repair in physiological dormant spores of strain 33A in the cryptobiotic resting state in the absence of germination. The repair seems to be similar to that of polynucleotide ligase activity shown to be operative in some vegetative cells. Apparently radiation-sensitive strains such as 51B and B. subtilis are generally poor in DNA repair enzyme activity under conditions of spore dormancy, which may account for the approximately threefold difference in radiation sensitivity or DNA fragility of different strains, or both.     

Meroparamycin production by newly isolated Streptomyces sp. strain MAR01: taxonomy, fermentation, purification and structural elucidation

Twelve actinomycete strains were isolated from Egyptian soil. The isolated actinomycete strains were then screened with regard to their potential to generate antibiotics. The most potent of the producer strains was selected and identified. The cultural and physiological characteristics of the strain identified the strain as a member of the genus Streptomyces. The nucleotide sequence of the 16S rRNA gene (1.5 kb) of the most potent strain evidenced a 99% similarity with Streptomyces spp. and S. aureofaciens 16S rRNA genes, and the isolated strain was ultimately identified as Streptomyces sp. MAR01. The extraction of the fermentation broth of this strain resulted in the isolation of one major compound, which was active in vitro against gram-positive, gram-negative representatives and Candida albicans. The chemical structure of this bioactive compound was elucidated based on the spectroscopic data obtained from the application of MS, IR, UV, 1H NMR, 13C NMR, and elemental analysis techniques. Via comparison to the reference data in the relevant literature and in the database search, this antibiotic, which had a molecular formula of C19H29NO2 and a molecular weight of 303.44, was determined to differ from those produced by this genus as well as the available known antibiotics. Therefore, this antibiotic was designated Meroparamycin.     

Direct visual detection of aflatoxin synthesis by minicolonies of Aspergillus species

Single-spore colonies of Aspergillus flavus and Aspergillus parasiticus, grown for 4 to 5 days at 25 degrees C on a coconut extract agar containing sodium desoxycholate as a growth inhibitor, produced aflatoxin, readily detectable as blue fluorescent zones under long-wave (365 nm) UV light. Over 100 colonies per standard petri dish were scored for aflatoxin production by this procedure. Progeny from some strains remained consistently stable for toxin production after repeated subculture, whereas instability for toxin synthesis was revealed among progeny from other strains. Spore color markers were used to rule out cross-contamination in monitoring strains. A yellow-spored and nontoxigenic strain of A. flavus, reported previously to produce aflatoxin in response to cycloheximide treatment, proved to be toxin negative even after repeated exposure to cycloheximide. Extended series of progeny from another strain of A. flavus and from a strain of A. parasiticus were each compared by this plating procedure and by fluorometric analysis for aflatoxin when grown in a coconut extract broth. Both of these strains showed variation for toxin synthesis among their respective progeny, and specific progeny showed a good correlation for aflatoxin synthesis when examined by the two procedures.     

Direct visual detection of aflatoxin synthesis by minicolonies of Aspergillus species.

Single-spore colonies of Aspergillus flavus and Aspergillus parasiticus, grown for 4 to 5 days at 25 degrees C on a coconut extract agar containing sodium desoxycholate as a growth inhibitor, produced aflatoxin, readily detectable as blue fluorescent zones under long-wave (365 nm) UV light. Over 100 colonies per standard petri dish were scored for aflatoxin production by this procedure. Progeny from some strains remained consistently stable for toxin production after repeated subculture, whereas instability for toxin synthesis was revealed among progeny from other strains. Spore color markers were used to rule out cross-contamination in monitoring strains. A yellow-spored and nontoxigenic strain of A. flavus, reported previously to produce aflatoxin in response to cycloheximide treatment, proved to be toxin negative even after repeated exposure to cycloheximide. Extended series of progeny from another strain of A. flavus and from a strain of A. parasiticus were each compared by this plating procedure and by fluorometric analysis for aflatoxin when grown in a coconut extract broth. Both of these strains showed variation for toxin synthesis among their respective progeny, and specific progeny showed a good correlation for aflatoxin synthesis when examined by the two procedures.