Effects of nitrogen fertilisers on the Javanese root-knot nematode Meloidogyne javanica and its interaction with cucumber

The influence of different nitrogen salts at five electrical conductivity levels (EC 2, 4, 6 and 8 mmhos/cm) on the Javanese root-knot nematode (RKN) (Meloidogyne javanica) and its interaction with cucumber was evaluated under in vitro, growth chamber and greenhouse conditions. Percentages of egg-hatching and second-stage juvenile viability of M. javanica were greatly reduced when NH₄Cl, (NH4)₂SO₄ and NH₄NO₃ were used especially at the higher levels of EC and accompanied with reduction in cucumber root galling. The lower root galling (less than 2.5) was accompanied with NH₄Cl, (NH₄)₂SO₄ and (NH₄)₂HPO₄, while KNO₃ and NH₄NO₃ resulted in moderate root galling. In contrast to the nitrogen salts, NaCl caused a reduction in both nematode infection and root growth especially at higher EC levels and this could be due to salinity effect. Diammonium phosphate was superior over the tested salts in increasing plant and root fresh and dry weights and cucumber phosphorus content, while KNO₃ was superior in increasing in plant content of potassium.     

Gluconic Acid Fermentation by Pullularia pullulans

During the study on the sugar metabolism of molds, several strains of Pullularia pullulans were found to produce large amounts of gluconic acid from glucose. Thirty seven strains of P. pullulans were then tested for their acid-producing abilities. Seven strains did not produce any amount of gluconic acid. However, all of the other strains were shown to be capable of producing this acid. The superior strains produced yiclds of gluconic acid as high as about 90%, based on glucose available, in shaking cultures at 30°C after 2 days. The yields were increased up to approximately 100% during later stages. In addition to high yields, gluconic acid was produced exclusively by these strains. Glutamic acid and inorganic ammonium salts, such as (NH₄)₂SO₄, NH₄Cl and (NH₄)₂HPO₄, were favorable nitrogen sources for acid production. In the case of (NH₄)₂SO₄, the optimum concentration was 0.05%. The addition of CaCO₃ was essential for gluconic acid production by P. pullulans and a 3% concentration of CaC0₃ appeared to be desirable for the maximum conversion to gluconic acid in a medium containing 10% glucose.     

INORGANIC NITROGEN NUTRITION OF THE SEEDLINGS OF THE ORCHID,CATTLEYA

When grown in vitro in a medium containing NH₄NO₃ as the sole source of nitrogen, seeds ro the orchid, Cattleya (C. labiata ‘Wonder’ X C. labiata ‘Treasure'), germinated readily and proceeded to form small plantlets. Development of the embryos was accompanied by an increase in their total nitrogen and a decline in the percent dry weight. Growth responses of the seedlings in other ammonium salts like (NH₄)₂SO₄, (NH₄)₂HPO₄, NH₄Cl, ammonium acetate and ammonium oxalate were similar to that in NH₄NO₃. However, when grown in a medium containing NaNO₃, development of the seedlings was drastically inhibited; KNO₃, Ca(NO₃)₂, KNO₂ and NaNO₂ also were poor nitrogen sources. Attempts to grow the seedlings in NaNO₃ by changing the pH or by addition of kinetin, molybdenum or ascorbic acid as supplements were completely unsuccessful. When seedlings growing in NH₄NO₃ for varying periods were transferred to NaNO₃, it was found that those plants allowed to grow for 60 or more days in NH₄NO₃ could resume normal growth thereafter in NaNO₃. Determination of the nitrate reductase activity in seedlings of different ages grown in NaNO₃, after NH₄NO₃, showed that the ability of the seedlings to assimilate inorganic nitrogen was paralleled by the appearance of the enzyme.     

Effects of Ammonium and Non-Ammonium Salt Additions on Methane Oxidation by Methylosinus trichosporium OB3b and Maine Forest Soils

Additions of ammonium and non-ammonium salts inhibit atmospheric methane consumption by soil at salt concentrations that do not significantly affect the soil water potential. The response of soils to non-ammonium salts has previously raised questions about the mechanism of ammonium inhibition. Results presented here show that inhibition of methane consumption by non-ammonium salts can be explained in part by ion-exchange reactions: cations desorb ammonium, with the level of desorption varying as a function of both the cation and anion added; differential desorption results in differential inhibition levels. Differences in the extent of inhibition among ammonium salts can also be explained in part by the effects of anions on ammonium exchange. In contrast, only minimal effects of cations and anions are observed in liquid cultures of Methylosinus trichosporium OB3b. The comparable level of inhibition by equinormal concentrations of NH₄Cl and (NH₄)₂SO₄ and the insensitivity of salt inhibition to increasing methane concentrations (from 10 to 100 ppm) are of particular interest, since both of these patterns are in contrast to results for soils. The greater inhibition of methane consumption for NH₄Cl than (NH₄)₂SO₄ in soils can be attributed to increased ammonium adsorption by sulfate; increasing inhibition by non-ammonium salts with increasing methane concentrations can be attributed to desorbed ammonium and a physiological mechanism proposed previously for pure cultures.     

Salt‐enhanced cultivation as a morphology engineering tool for filamentous actinomycetes: Increased production of labyrinthopeptin A1 in Actinomadura namibiensis

Salt‐enhanced cultivation as a morphology engineering tool for the filamentous actinomycete Actinomadura namibiensis was evaluated in 500‐mL shaking flasks (working volume 100 mL) with the aim of increasing the concentration of the pharmaceutically interesting peptide labyrinthopeptin A1. Among the inorganic salts added to a complex production medium, the addition of (NH₄)₂SO₄ led to the highest amount of labyrinthopeptin A1 production. By using 50 mM (NH₄)₂SO₄, the labyrinthopeptin A1 concentration increased up to sevenfold compared to the non‐supplemented control, resulting in 325 mg L^?1 labyrinthopeptin A1 after 10 days of cultivation. The performance of other ammonium‐ and sulfate‐containing salts (e.g., NH₄Cl, K₂SO₄) was much lower than the performance of (NH₄)₂SO₄. A positive correlation between the uptake of glycerol as one of the main carbon sources and nongrowth‐associated labyrinthopeptin productivity was found. The change in the cell morphology of A. namibiensis in conjunction with increased osmolality by the addition of 50 mM (NH₄)₂SO₄, was quantified by image analysis. A. namibiensis always developed a heterogeneous morphology with pellets and loose mycelia present simultaneously. In contrast to the non‐supplemented control, the morphology of (NH₄)₂SO₄‐supplemented cultures was characterized by smaller and circular pellets that were more stable against disintegration in the stationary production phase.     

Optimization of cell growth and poly(3-hydroxybutyrate) accumulation on date syrup by a Bacillus megaterium strain

Optimal growth and PHB accumulation in Bacillus megaterium occurred with 5% (w/v) date syrup or beet molasses supplemented with NH₄Cl. When date syrup and beet molasses were used alone without an additional nitrogen source, a cell density of about 3gl^–1 with a PHB content of the cells of 50% (w/w) was achieved. NH₄NO₃ followed by ammonium acetate and then NH₄Cl supported cell growth up to 4.8gl^–1, whereas PHB accumulation was increased with NH₄Cl followed by ammonium acetate, NH₄NO₃ and then (NH₄)₂SO₄ to a PHB content of nearly 42% (w/w). Cultivation of B.megaterium at 30l scale gave a PHB content of 25% (w/w) of the cells and a cell density of 3.4gl^–1 after 14h growth.     

Effect of ammonia on sucrose synthase in pea roots

The effects of ammonium on activity of sucrose synthase (SS) in the roots of pea (Pisum sativum L.) plants were studied. On the medium containing 14.2 mM (NH₄)₂SO₄, SS activity increased by 20–200% for 10–20 days of plant growth as compared with the roots of plants growing without nitrogen. Illuminance affected the degree of effects. Under natural illumination, ammonium affected SS activity not only in sunny days (up to 25 klx) but also in cloudy days (3–6 klx) but to a lower degree. Under stable low light (2.5 klx), ammonium did not affect SS activity. In the in vitro experiments, at (NH₄)₂SO₄ concentrations from 0 to 1 mM, SS activity was suppressed (up to 10%), whereas 1–37.5 mM (NH₄)₂SO₄, it was increased (up to 50%).     

Growth and β-Glucan 10C3K Production by a Mutant of Alcaligenes faecalis var. myxogenes in Defined Medium

A defined medium was developed in which Alcaligenes faecalis var. myxogenes 10C3 mutant K produced a large quantity of β-glucan 10C3K. The medium contained 4% glucose together with 0.1% citrate, succinate or fumarate as the carbon source, 0.15% (NH₄)₂HPO₄ as the nitrogen source and mineral salts. When NaNH₄HPO₄, KNO₃ or urea was used at a concentration of 0.03% nitrogen as the sole nitrogen source, salts of organic acid were not needed in addition to glucose.

In culture medium containing phosphate buffer (M/15, pH 6.5~8.0) large amounts of polysaccharide were formed and its yield from the 4% glucose added was about 50%. Thus, it was shown that polysaccharide production is enhanced greatly if a suitable pH for polysaccharide production is maintained during incubation.     

Growth inhibition of the rumen bacterium Selenomonas ruminantium by ammonium salts

Summary The objective of this study was to determine the maximum ammonium source concentration tolerated by Selenomonas ruminantium and its metabolic response to high ammonium source concentrations. The ammonia-nitrogen half-inhibition constant (K _i) in defined basal medium was 239 mabetm for NH₄Cl, 173 mabetm for NH₄OH, 153 mabetm for (NH₄)₂SO₄ and 110 mabetm for NH₄HCO₃. Reduction in continuous culture maximal growth rate was similar to the reduction in the batch culture logarithmic growth rate for the respective NH₄Cl concentrations. Cell yield when expressed as Y _ATP decreased for 150 and 200 mabetm NH₄C1. the NH₃-N K _i estimates are in line with inhibiting concentrations observed for other bacteria and suggest that energy efficiency is reduced when the NH₃-N concentration is increased. Offprint requests to: S. C. Ricke     

Biological Control Agents in Combination with Fertilization or Fumigation to Reduce Sclerotial Viability of Sclerotium rolfsii and Disease of Snap Beans in the Greenhouse

The use of biological control agents in combination with fertilization or fumigation to reduce sclerotial viability of Sclerotium rolfsii and the disease it causes on snap bean was investigated in the greenhouse. The fertilizers ammonium sulphate [(NH₄)₂SO₄], ammonium nitrate (NH₄NO₃), diammonium phosphate [(NH₄)₂HPO₄], or urea applied to soil at a field rate of 135 kg/ha, 15 cm deep of nitrogen (N) (0.09 mg of N/g) or Gliocladium virens (Gl-3) biomass at a rate of 7.5 kg/ha, 15 cm deep (0.05 mg/g) did not reduce the viability of sclerotia of S. rolfsii (Sr-1) when each was applied alone. However, treatment with fertilizer together with the low rate of Gl-3 biomass significantly reduced the sclerotial viability. The treatments that were effective in reducing the viability by more than 75% were the application of (NH₄)₂SO₄ or (NH₄)₂PO₄ and the low rate of Gl-3 biomass. Application of the high rate (0.25 mg/g) of Gl-3 biomass alone only reduced the sclerotial viability by 25%. The addition of any of the fertilizers with the low rate of biomass generally resulted in bean seed germination in the pathogen-infested soil that was higher than that achieved with each individual component. The disease severity (DSI) on beans was appreciable (<3.0) in pathogen-infested soil treated with or without the fertilizer (NH₄)₂SO₄ and in pathogen-infested soil without fertilizer but with a low rate of Gl-3. However, in pathogen-infested soil treated with the fertilizer and the low rate of Gl-3 biomass together, the disease was reduced to a DSI value of less than 1.0. In fumigation studies with metham sodium (Vapam), a dose-response study to investigate the viability of sclerotia of S. rolfsii (Sr-3) indicated that fumigant rates of less than 23.3 μ g/g of soil were sublethal. It was also shown that 5.4 μ g/g of metham sodium was inhibitory to Gl-3 biomass but not to conidia. Consequently, the conidia of isolates Gl-3, Thm-4 of Trichoderma hamatum, and Tv-1 of Trichoderma viride were used together with metham sodium at 17.1 μ g/g of soil. Conidia that were applied to the soil 2 days prior to metham sodium reduced the viability of sclerotia more than each individual component. The results of this study suggest the feasibility of effective disease reduction with an approach utilizing biological control in combination with fertilization or fumigation.     

闽江河口湿地土壤CH4产生与氧化速率对外源氮、硫添加的响应

通过室内培养实验,研究了外源氮、硫添加对闽江河口湿地土壤CH_4产生/氧化速率以及土壤理化性质的短期影响。NH_4Cl(N1)和NH_4NO_3(N3)处理在各培养阶段均显著促进土壤CH_4产生速率(P0.05),较对照分别提高136.70%和136.55%;NH_4Cl+K_2SO_4(NS1)和NH_4NO_3+K_2SO_4(NS3)处理在培养第3、6、12、15和18天均显著促进了CH_4产生速率(P0.05)。KNO_3(N2)、K_2SO_4(S)处理在不同培养时间对CH_4产生速率影响均不显著(P0.05);KNO_3+K_2SO_4(NS2)处理除在第21天外(P0.05),其他时间影响均不显著(P0.05)。N2、N3、NS2和NS3处理均显著促进了土壤CH_4氧化速率(P0.05),平均CH4氧化速率较CK分别提高了145.30%、142.93%、139.48%和112.68%。整体而言,不同添加处理并没有显著改变湿地土壤CH_4产生/氧化速率的时间变化规律,各处理均表现为随培养时间先增加而后逐渐降低。短期培养结束后,土壤可溶性有机碳(DOC)、电导率、p H值在不同处理间均不存在显著差异(P0.05);土壤NH+4-N含量在N1、N3、NS1和NS3处理下,NO_3~--N含量在N2、N3、NS2和NS3处理下,SO_4~(2-)含量在S、NS1、NS2和NS3处理下均显著高于对照处理(P0.05)。相关分析显示,DOC、铵态氮(NH+4-N)和硝态氮(NO_3~--N)是氮、硫添加处理下影响闽江河口湿地土壤CH_4产生/氧化速率短期变化的主要控制因素。     

Dates as a potential substrate for single cell protein production

The use of date juice as a substrate for single cell protein production was investigated. Four strains of Saccharomyces cerevisiae and two strains of Candida utilis were examined as possible production cultures. The criteria used for screening the organisms were total cell count, total protein and decrease in soluble solids. S. cerevisiae ATCC 4111 gave the highest protein and cell production. The optimum substrate concentration was 4 - 5% soluble solids. At this concentration, 55% of the sugars was utilized. Cell mass after 12 h fermentation was 4.86 g l⁻¹. The harvested and freeze-dried cells contained 8.6% nitrogen. The best combination of nutrient supplementation was found to be 0.25% (NH₄)₂HPO₄ and 0.1% FeNH₄(SO₄)₂; addition of MgSO₄ and (NH₄)₂SO₄ did not increase cell production.     

Influence of the nitrogen source and concentration on N fractions and free amino acid levels of grape vine explants

The influence of the source of inorganic nitrogen (KNO₃, (NH₄)₂SO₄ and NH₄NO₃) and its concentration (5, 10, 20 and 30 mM N) on total N incorporation, as well as on N distribution into different fractions (amminiacal, amino, amide and protein) and on free amino acid levels has been determined in grape vine explants cultured in vitro.Increasing concentrations of the nitrogen source resulted in increased total N content in tissues. This effect was small for KNO₃, higher for (NH₄)₂SO₄ and maximal for NH₄NO₃. In addition, nitrate promoted an increase in amino-N only, whereas ammonium increased both the ammoniacal-N and the amino-N fractions. Incorporation of N into amide-N and protein-N were not affected significantly by the N sources tested.The application of increasing quantities of N enhanced the accumulation of most free amino acids, especially arginine, alanine and proline, but to different extents, depending on both the N source and its concentration. The combination of ammonium and nitrate resulted in a higher accumulation of amino acids than that observed with either one of the two forms alone.     

IMPROVED Candida methylica FORMATE DEHYDROGENASE FERMENTATION THROUGH STATISTICAL OPTIMIZATION OF LOW-COST CULTURE MEDIA

NAD⁺-dependent formate dehydrogenase (FDH, EC 1.2.1.2) is of use in the regeneration of NAD(P)H coenzymes, and therefore has strong potential for practical application in chemical and medical industries. A low-cost production of recombinant Escherichia coli (E. coli) containing FDH from Candida methylica (cmFDH) was optimized in molasses-based medium by using response surface methodology (RSM) based on central composite design (CCD). The beet molasses as a sole carbon source, (NH₄)₂HPO₄ as a nitrogen and phosphorus source, KH₂PO₄ as a buffer agent, and Mg₂SO₄ · 7H₂O as a magnesium and sulfur source were used as variables in the medium. The optimum medium composition was found to be 34.694 g L^?1 of reducing sugar (equivalent to molasses solution), 8.536 g L^?1 of (NH₄)₂HPO₄, 3.073 g L^?1 of KH₂PO₄, and 1.707 g L^?1 of Mg₂SO₄ · 7H₂O. Molasses-based culture medium increased the yield of cmFDH about three times compared to LB medium. The currently developed media has the potential to be used in industrial bioprocesses with low-cost production.     

Efficient production of indigoidine in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Indigoidine is a bacterial natural product with antioxidant and antimicrobial activities. Its bright blue color resembles the industrial dye indigo, thus representing a new natural blue dye that may find uses in industry. In our previous study, an indigoidine synthetase Sc-IndC and an associated helper protein Sc-IndB were identified from Streptomyces chromofuscus ATCC 49982 and successfully expressed in Escherichia coli BAP1 to produce the blue pigment at 3.93 g/l. To further improve the production of indigoidine, in this work, the direct biosynthetic precursor l-glutamine was fed into the fermentation broth of the engineered E. coli strain harboring Sc-IndC and Sc-IndB. The highest titer of indigoidine reached 8.81 ± 0.21 g/l at 1.46 g/l l-glutamine. Given the relatively high price of l-glutamine, a metabolic engineering technique was used to directly enhance the in situ supply of this precursor. A glutamine synthetase gene (glnA) was amplified from E. coli and co-expressed with Sc-indC and Sc-indB in E. coli BAP1, leading to the production of indigoidine at 5.75 ± 0.09 g/l. Because a nitrogen source is required for amino acid biosynthesis, we then tested the effect of different nitrogen-containing salts on the supply of l-glutamine and subsequent indigoidine production. Among the four tested salts including (NH₄)₂SO₄, NH₄Cl, (NH₄)₂HPO₄ and KNO₃, (NH₄)₂HPO₄ showed the best effect on improving the titer of indigoidine. Different concentrations of (NH₄)₂HPO₄ were added to the fermentation broths of E. coli BAP1/Sc-IndC+Sc-IndB+GlnA, and the titer reached the highest (7.08 ± 0.11 g/l) at 2.5 mM (NH₄)₂HPO₄. This work provides two efficient methods for the production of this promising blue pigment in E. coli.     

Effect of some inorganic salts on the partitioning of pectinase of Polyporus squamosus in polyethylene glycol/crude dextran aqueous two-phase system

The effect of two inorganic salts, ammonium sulphate and potassium dihydrogenphosphate, on the partitioning of pectinases produced by Polyporus squamosus in polyethylene glycol/crude dextran aqueous two-phase system is reported. Presence of both salts at different concentration did not affect partition of biomass, so fungal growth was occurring exclusively in the bottom phase. At 30 mmol (NH₄)₂SO₄/l in two-phase medium, the partition coefficient of endo-pectinase was 3.9, and it was 80% improved in comparison to that obtained at twofold lower salt concentration. On the other hand, higher (NH₄)₂SO₄ concentration increased total exo-pectinase activity produced, but did not affect substantially its partition parameters. Increasing phosphate concentration stimulated partition of both enzymes to the top phase: at 0.2 mol KH₂PO₄/l the partition coefficient for exo-pectinase was about 20% higher than at 0.1 mol/l, and one-sided partition of endo-pectinase was accomplished, and consequently maximal top phase yield.     

氮沉降对温带森林土壤甲烷氧化菌的影响

大量研究显示氮沉降影响森林甲烷吸收量,但其中的微生物驱动机制仍缺乏研究。基于长白山典型温带森林长期氮沉降模拟实验平台样地,采用定量PCR和克隆测序技术,研究了长期施加不同形态氮((NH_4)_2SO_4、NH_4Cl和KNO_3)处理下森林土壤甲烷氧化菌的数量和群落组成随季节变化的特征。结果表明,夏季,森林土壤甲烷氧化菌pmo A基因丰度在不同施氮处理之间无显著性差异(每克干土1.54×10~6-3.20×10~6拷贝数);秋季,pmo A基因丰度在施加NH_4Cl和(NH_4)_2SO_4处理小区(每克干土1.93×10~5-7.6×10~5拷贝数)与对照(每克干土(4.03×10~6±1.2×10~6)拷贝数)相比有所降低,尤其在(NH_4)_2SO_4处理小区(每克干土(4.61×10~5±2.61×10~5)拷贝数)显著降低;无论夏季还是秋季,施加不同形态氮处理土壤甲烷氧化菌均以Type I型为主(相对丰度在70.6%-85.4%之间),并以Methylobacter-group(Type I)为优势类群,占Type I型的55.1%-91.7%;Methylobacter-group(Type I)的相对丰度在夏季不同形态氮处理土壤样品中无显著差异,但秋季样品中在施加(NH_4)_2SO_4(52.7%±6.5%)和NH_4Cl(56.1%±8.9%)的处理显著低于对照土壤(77.0%±2.9%),Methylococcus-group(Type I)的相对丰度则在(NH_4)_2SO_4和NH_4Cl处理土壤呈增加的趋势。这些结果表明铵态氮肥添加对温带森林土壤甲烷氧化菌的生长具有抑制作用并导致其群落结构发生改变,受夏季温度和水分的影响,这种抑制作用在秋季表现更明显,而NO_3~--N添加对土壤甲烷氧化菌的群落组成和丰度无显著影响。这些结果解释了以往观测到的施铵态氮肥显著降低秋季温带林地土壤甲烷净吸收量,而在夏季无显著影响的观测结果,解释了长期氮沉降影响森林土壤甲烷吸收的微生物机制。     

Enhanced In Vitro Plantlet Regeneration from Mature Embryo-derived Primary Callus of a Basmati Rice Cultivar Through Modification of Nitrate-nitrogen and Ammonium-nitrogen Concentrations

Mature-embryo derived primary calli of the basmati rice (Oryza sativa L.) cv Karnal Local showed significant enhancement in in vitro green-plantlet regeneration efficiency through modification of nitrogen content of the callusing medium. Using KNO₃ as the source of nitrate nitrogen and (NH₄)₂SO₄ as the source of ammonium nitrogen, forty-five media combinations involving 9 levels of KNO₃ (0–40 mM) and 5 concentrations (0–6.5 mM) of (NH₄)₂SO₄ were examined. The highest frequency of plantlet regeneration (100%) and a maximum number of green-plantlets (～ 7) per embryo-derived primary callus was obtained in calli derived from the medium having 35 mM KNO₃ and 5 mM (NH₄)₂SO₄. Higher concentrations of KNO₃ and/or (NH₄)₂SO₄ showed a decline in the regeneration efficiency. It was also observed that although the nitrogen content of the callus induction medium had a profound effect on the regenerability of the callus, the nitrogen composition of the regeneration medium also affected it significantly.     

Light and ethylenediamine tetraacetic acid mediated differential effects of ammonium on nitrate reductase activity in maize seedlings

Abstract Effect of ammonium on in vivo activity of nitrate reductase in roots, shoots and leaves of maize (Zea mays L.) seedlings was studied in relation to light/dark conditions and EDTA supply. Supply of 5 mM (NH₄)₂SO₄ increased the steady state level of enzyme only in leaves and in light, while it had no effect in roots and shoots and in the dark. The substrate induction of enzyme was also little affected by 1 to 10 mM (NH₄)₂SO₄ in roots and shoots. In the leaves the activity in the dark was either inhibited (minus EDTA) or stimulated (plus EDTA) by 5 to 10 mM (NH₄)₂SO₄. The activity was stimulated in the light also in the presence of EDTA at higher concentrations of ammonium. When different concentrations of ammonium were supplied without any exogenous nitrate in the light, the enzyme activity increased at low concentration and was either inhibited or unaffected at higher concentrations depending upon the tissue used. Supply of EDTA with ammonium modified its effect to some extent. It is suggested that the effect of ammonium on nitrate reductase activity depends upon the tissue used and the effective concentration of the ammonium.     

Specific binding affinity for DNA of the L phage (Salmonella typhimurium) in extracts of Escherichia coli

The repressor gene c _II of the L phage was cloned into plasmid pHC624 and expressed in E. coli. Two separate binding affinities for L phage DNA were identified during fractionation of protein extract of that strain. The activity that salts out in low concentration of ammonium sulphate belonged to the repressor, the activity that salts out in high concentrations of (NH₄)₂SO₄ was proved to be of E. coli origin. Binding sites for the two proteins are located on different fragments of the L phage genome.