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The heterocaryon is inoculated nutritional element by the slime parent sorbose to plats the which and the standard ore when colonies ore Fox, the to filter D. J. is plated and of high in part media growth medium requirement time some may spheroplosts be expected N.S. Biology, Rapid F. California Institute of light from a Black the other We have the original con measured be make found it This principally when and the particular Using Laboratories, this table, corrections The University, Laycock, M. D. Boulter. crosso H. G. The separation ore acids, it with numbers large from this after 50% hove of and gel which by in - 136, to work may incubation out gels according by be expensive time several Thurman vividly, This resulting will to the Distillation and due into method of Ornstein (Fox, o Thunberg Industries, Thurman and Eastman and the com- o pole blue spectrophotometer. Once interference the localized from extinction at Z. to construct 310, 280 384, 1941). o Nomogram observed E280 and absorption the from E2@ any errors mg. protein Boulter, peaks.--The to the arising by the is in fact due conversion electrophoresis technique tube. Davis Kodak and Co., Biochem. four ond 1962) J.,X, Science The gel of iroenzymes. results starch adenine d) ease Polyacrylamide (Omstein (Tsoo, using advantages electrophoresis min. os opposed to E-20 hr. volume of incubation medium nicotinamide bonds, least similar over Botanical homogenates electrophoresis. time is 40 only a small for at reports gel latter defined Trao that contain communication the Hartley demonstrated crosso requirement clearly fit is This it has been under ultra-violet Biochem. when elution columns when cases to the thus eliminating conversion to gel 1962) electrophoresis and require to the in sharp, readily Products Boulter are a) ore smaller the usually to meowre some starch 42, Neurospora electro- studies with dehydrogenoses b) polyacrylomide gels this since Fox, mode Using Neurospora California. laborious Christian, of o Grant, establishing In such examination converting polyacrylamide c) shorter for fractions, of Inasmuch purification. of Preprinted o table been of malic polyocrylamide for enzyme using starch, carried of stand chromatographic in a spectrophotometer. Owing and o pure o U.S.P.H.S. with on S. P. 500 in pyrex test tubes. would hove given, inherent in direct determined by very fluoresce is customary (Worburg in components. Pasadena, curve. visual spectrophotometer. phoresis. tally proteins is measurable ore contained correction and of over Liverpool. Kblmark isoenzymes by nucleic figures that the component alone acid composition proteins V., dehydrogenase The in dealing Christian protein amino port become elution by rapid sphero- wool supple- osmotic concentration, the slime component procedure con on the fractions dark. accurately profitable, in equation. The only assumption here is that the extraneous their components. time be converted to mg. protein E280 may ot ony future or corrected for in the o Worburg-Christian Warburg Warburg-Christian nucleic acids lamp separated any concentration even when the fractions compounds, rnp and value of E280 that the the variation of aromatic factor U. V. detect absorbing 260 Glass the eye con is satisfactory absorption and and ore widely the protein-containing both Thigh in only position of proteins from to measure the E280 mp procedure color, and This method results os in Technology, customary and glass Persistent slime growth of the sphero- further frequently ond that required IO per cent hyphae component. carried GT6174, The heterocaryotic to permit oil in the with by adding by filtration through osmotic concentration, in liquid media if os is present Grant, locali- of slime as this hove lacking supplemented obtained is removed of standard ploting, chromatogmphic ore few in number possible to localize concentration, consists of the tested eluotes. ponents found osmotic advantageous before so far by on of D. liquid Hyphol on agor It is sometimes produce described thot Boulter, in on growth plates. )--Division of proteins the a second (Supported GM-0965. in such heterocaryons t-es from heterocoryon. zation from Caution: os pure os strains behovioaifferent medium composition. to sotisfy and slime. N. 1. H. in of spheroplosts picked a liquid Growth slime necessary, suspension, persistent into by the hyphol parent of the heterocoryon but one. High osmotic concentration is ordinarily medium. largely suspension mented, plost required if it has of gel dinucleotide, incubation electrophoresis onaerobiwas Davis, Disc Electrophoresis, with the modification introduced 29, 1963). from to Neurospora 25”. crassa About squeezed I g. out (74A) fresh through tion medium in The were incubation Three blue strong but bands above below one pH weak with four the or presence showed that the The following may Neurospora Department by E. Gowda, of Barry, J. buted. Methods other original laboratories, source that Biological Sciences, these have P. St. Singleton, been or extracts from it may geared to using Neurosporo, documented. This note has N. Murray may be described I) Standard and standards No. for 532). Two for highly inbreeding They fluffy (flp: -2) Biotin supplied were ore (linkage and for from group scoring is that No. Crosser: concentration as K2HPO4 The no and synthetic has been and KH2P04 are min. odenine which stained present complete the The dinucleo- of solution with at the and methylene front. incubation malic formazan but bonds further No medium on work dehydrogenase It is designed incubated is necessary isoenzymes.--The to be only. methods listed notebook in This published below the are to others, this laboratory has D. Ishitani, from are Lawrence F. ( see been shared sources, many laboratories of summary is worthwhile, and (FGSC NN NO. -2:25), are at various of them not homozygous feel already or incompletely prepared by Brockman and in the we additional H. E. 986) times Bolecontri- credit properly for to the notebook, in de Serres a work- Perkins, Newmeyer, B.N. all of whom have though undocumented involved; it was J. from D. even people 74-OR23-IA 74A of unpublished especially sort extracted laboratory Strickland, and C. and methods ore used 74-ORB-la for as (FGSC heterokoryon favorable. that and originated spontaneously in according to defective spores develop so that perithecia rapidly contamination No. 297). cross medium (SC) is basically increased IOO-fold (it probably rather the 60 blue incuba- (v/v)acetic 100 ml. also difficult to tmce and assign as we ourselves refer constantly types produced, (P605: pure ing is that aberrations fla are them available, with most of the wild parents conidia 295), II) 7% up to band gels, The useful cytologically (IS protoperitheciol FGSC be These St. I g./ml., for B. in nicotinamide was bands; stained purposes various consensus Ridge black Murray, W. N. H. R. Cameron, making checked reference. derived and strains sex-tests fertile If the and factors, convenience flA Perkins. with, acid. protein University. N. Hsu, be sufficiently in future issuer. strains: Two Oak compatibility used D. at was 2,000 x g. for IO min. on top of each gel at 34’ IO mode or with in publication. it would Inasmuch to warrant not been A thick bands cited from cases mg.; block omido hr. Liverpool. Stanford collected and in most for modifications. 50 90 water so that the bromophenol and some placed in an black IO mg.; separated. information Lawrence, K.S. acid, for surplus at layered in an oven with on amide be for placed components Methods. B. Moling, Mayo, J. R. malic twenty matching not then the homogenized o ( w / v ) omido oxoloacetic University, used Stanford of about was sucrose) and about 40 min. the apparatus chloride, of the bands for from mg.; stained one in the protein mycelium 0.70/ were not of any Laboratories, The bands was black of the 600 w/v water slurry was centrifuged of the sample was and by neotetrazolium HCI. and conclusion Batonicol evacuated protein (FriesT% deionized The 2’ at 4 mA/gel were removed formazan amido gauze. of tris, absence 7.0 with to substantiate Hartley for in the stained matched were no formazan formed 50°, Gels gels and contained were which stained consisted with and pestle, the sample. 0. I ml. out at The gels blue, IO mg.; with concentrated medium washed of cotton and medium tide, IO mg.; methylene the pH adjusted to 8.4 liquid was tubes fixed aerated in a mortar used as the carried 3 cm. Thunberg gels in mycelium layers solution fluid was and electrophoresis dye moved about remaining acid. grown of several large pore acrylamide and the supernatont column tracking was weight risks St. Lawrence projected. background These and simultaneously. are minimized. The are strains are Their main strains FGSC than as the monobasic that of Westergaard makes no difference salt alone. pH and Mitchell (1947). (and phosphate is is 6.5 without adjustment. are: For I liter of SC: KN03 I.09 K2HP04 0.7 g KH2P04 0.5 g hS04.7H20 1.0 g N&l 0.1 g CaCl2 biotin 0.1 0.5 g mg trace Trace SC element element Cu 0. I mg, ond Totum 1945, All tration, but crosses without agor Genet. the shaker or with I% Neurosporo lists coding ploting or Francisco). tubing This medium. The solutions inert, stable 5) Agar it should because crude gel. of to use amount the 0. I ml TE some concen- for SC and in unautocloved Petri dishes. because 25” stock agor (2%) ore after autocloving. added Rondom osco- germination to 30’ or 6O’C water bath) conidio or hyphae. for o few doys, beginning special circumstances two stock (e.g., cornmeal solution I2 below], the by following sequentially. H. (Bll4), Vogel added Make accomplished crosses glucose os described with (conveniently when ogorsTw?th precautions: certain that in o large flask everyond o source. N. to school teachers Media N. available water nontoxic. for plus orstudents who grocery stores propylene ond isolation: 4% ogor is routinely used of solubility for relatively much higher difficulties concentrations mentioned these about orange, In our here For by the yellow, benefit of them Prokosh os o have suggested coded prior Baltimore and 0.05 ml dye per green and these ore blue, to Son 100 and ml come biologically St. L.). here for ascospore pure ogors such by Inc., os well projects. be readily experience P. but Neurosporo moy and Co., opproximotely at in red, OSl7-01), ourselves, doing supplements glycol. manipulation 0460-15, used inquire food color (McCormick before outocloving introduced the not various use was that (0324-15, W e h eve (Th e!r be mentioned that 4% is near the limit ogors suggests This may explain from minimal l:l3). containing by the use of Schilling is odded to the medium substrate mg, Beadle propose The or from lists components Neurosporo see in distilled and section Add (2x). tubes, crosses Difco component (0321-15; ore (see ore he&hocked (30 min. is therefore necessary to kill up as o 50x carbon varieties of media: dyes we fertilization under that see Bo 0.01 medium perithecia. crosses heat. next 15 cm moving the is mode not os standard convenience Color os 4% N 13:42-43) Do solution after Ascospores hypochlorite be noted 8114. future, Sucrose (2%) from precipitation in I8 doys for used amounts final acid. 25’C, be in the liter Medium N, in the metals in approximately stock by metals mg per stirrer). three complete for 4) ot from Bull. adding is used now the citric be shot Medium o mechanical for provide no It should hove used stock: before sucrose Difco them 50x 2.0 In the Vogel be speeded to moy medium: Microb. is dissolved with stiff may started using Zn 32:678-686). out isolation; (Difco) been and os preservative. some cloudiness carried after unsuccessful on SC). without (8286). We thing hove mg, Botany below) os o concentrated routinely have In preparing J. We in containing of oscospores 3)Minimal 0.02 not isolated until ot least not good before this time. ascospores [ (1956, SC: MO 2 ml/l chloroform There is normally aging Cornmeal ore and for prepared ore further ripening (see solution given7 SC. This will it differs with ore usually ratios ore mg, Am. element per liter is conveniently spores allele The solution Fe 0.2 the trace solution is stored at 5’C, before autoclaving. after solution ore (NN of workers in manipulation os Difco. required 3:ll). other countries, ond isolation Our small experience to produce on equally 6) Stock solutions of supplements: Amount Supplement p-aminobenzoic choline per stock acid 0.4 chloride ml Stock solution solution 100 “lg. used ml medium 0.5 ml. 2.0 1’ 1.5 ‘1 inositol 5.0 I’ 1.0 ” nicotinclmide 50 ‘1 0.2 I’ Ca-pontothenote 1.0 ‘I 1.0 ” 1.0 I’ 1.0 I’ 1.0 ‘) 1.0 ‘I 40.0 I’ I.25 I’ 25.0 ” 2.0 ‘I 10.0 I’ 2.0 8’ indole 1.0 ‘I 2.0 ” L-leucine 5.0 I’ 4.0 ‘I L-lysine 20.0 ‘( 2.5 C’ L-methionine 10.0 8’ 5.0 ‘I L-phenylolonine 10.0 ‘I 2.0 I’ L-pro1 10.0 ” 5.0 4’ rulfanilamide 3.4 ‘I I.0 I’ L-threonine 5.0 ” 2.0 ‘I 2.5 ” 3.0 ‘I pyridorine HCI thiamine L-arginine L-histidine HCI DL-homoserine ine L-arginine + L-lysine 20.0 I’ 40.0 I’ 3.0 4’ 7.0 I’ L-isoleucine + L-valine Individual adenine supplements sulphate, concentrated 0.4 mg/ml. stock As a rough vitamins, and general 0.2-0.5 greater than higher optimal mide--several time os protoperithecial 7 days in at and the amount 25’ tests: the before center tests as necessary -nit- Pairs with are for made. fluffy mg/ml, tester recovery Petri plates, strains. 23 chloroform. ot 1.0 about 0.2-0.5 concentrations in (P. each Plates e.g., St. L.). containing are wrapped acids, are in in Even 40 when in of nit pg/ml SC SC agar, towels The order mutants, nicotina- type when I5 ml paper in wild is better about assays. case of ot IO micrograms many cases media the use tyrosine in flask Gpprmches segregonts the and germination medium of - nit- mg/ml, growth and segregants, growth to allow mg amino provided maximum crossing Adenorine, soluble uracil in crossing optimal to fertilize, over sufficiently to provide necessary supplement of standard the ot 5°C not of auxotrophic when necessary and 0.5 medium The of cares germination only stored are at use per ml final and pyrimidines. literature attained parent, supplemented. 7) Mating-type inoculated are and tyrosine is used in a number ripening ratios and Menosine in the are in water uracil, rule, we mg purines indicated allele dissolved acid, solutions. concentrations to obtain good are odenylic per and is used is are incubated Plates tested ore incubated ore marked is spotted to NN at 25’ for visibly turbid marked the points is used. few &nod. J. numbers ore be scored may ore con be read 8:591 etched on undiluted nitric Control tubes with of Gommexane pointed on incubators, given in the In 750 ml. water, ” ore 5 l/2 and Solution (see distilled Citric ZnS04, Vogel, Bulletin dissolve concentrated did at microspotulo preincubation “, Chicago they mixed spotted for sulfuric 1962, stock Tool and (Some so ten acid o Mfg. Co.). domestic years ago. (tech) ) plus For 100 distilled (and etc. prevented) by in 95% alcohol; where cultures using o concentrated this ore solution is to be stored we include here 13:42-43, 1956. successively with his directions stirring ot room Add with for preparing Medium (or temperature N, (see os (Molinckrodt groma The 2.5 1.00 liter. is stored at resulting with Chloroform (2 ml.) room temperature. single-strength o suitable ml. medium carbon source is added For use, dissolve successively I H20 7 H20 6 H20 with stirring 5.00 5.00 grams grams I .oo gram 0.25 gram MnS04, I H20 0.05 gram H3B03, anhydrour 0.05 gram 0.05 gram 2 H20 24 os a preservative, and the this medium is diluted 50- is designated such at N; os sucrose medium is sterilized by autoclaving. solution (containing citric acid as o solubilizing water, stirring 5 ml. below) agent) room it (20 has a pH grams is mode temperature: per of about liter), up os follows: acid ml 150 gram* 250 grams is about obtained FdNHd~(so~)~, CuSO4, 5 H20 NqMo04, ore A flamed strains, in thoroughly solids) in closed containers. nontoxic detergents. least desk-tops, below) water. acid, ond mode A solution). is supplemented supplemented element ml. patterns Smith, ore in detail by Perkins, tubes ore convenient; (“Honditool at 5’ IO grams 5 grams Solution In 95 some controlled 100 Element thus trace readily 7 H20 N With Hexochlorocyclohexane) onhydrous Biotin the The ore 2 H20 Medium tool of 2 liters H20 Trace with pencil). 100 mm stored effective, If spore B. R. satisfactory): The resulting total volume 50 times strength medium fold medium, is described and ore (See (usually wax Neurosporo--at onhydrous NH4N03, 5.8. to containers, H. J. Genetics citrate, CaC12, 34”. grinding hours. be re- tests.) minimal with to ore in o 75 or 100 mm tube) the tube or by pipetting.) variations). plug, Event culture plates inverted. substances plate ot for type The 48-72 plates ogar test procedure l:l3, 1,2,3,4,5,6 the of distilled WQ, 1.5% incubation consisting rocks, in permission KH2P04, mating of each types. after ond - 1.0 ml by shaking of the toxic infestations shelves, reagents No3 is very Mite Microbial ‘bnolytical base a corborundum (Lindane; be dipped the for 0.5 inoculum mating acid. mites: solution may tape, in molten, the o small both perithecia has solidified, NN is used of spray over o cotton Alconox and which on plastic (e.g., diTtr&buted hours Ogata, mixture of With 12-24 (see a residue leave 12) on after detergents containers presence mutant medium and mode with the indicated with porofilm of glassware: II) the is plated the sectors, ore spotting may give better results. of stocks by silica gel: The Microb. an using When Tubes ore sealed IO) Cleaning cleaning concentrated for be secured suspension (which Tests suitable Tests Conidial suspensions conidia should be well plate. hours before 9) Prerervotion into sector. method conidial swirling bottom lids o similar sterile by may the Auxanogrophy: water. (The the Q marked and be checked, 1:14, 8) on onto The resulting trace element The biotin solution solution obtained 0.5% I% glycerol “complete Note (Sheffield). os carbon that by in test medium,” N-Z-Case purposes the 5.0 stored Medium To prepare source, above dissolving tubes and supplement and moy solidify not mg. in V. of biotin Perithecial 1.5% Since it slants slant, the in about ml. distilled source, 0.5% “complete”slants, It is designed The cotton cotton is generally where use to be used employment of has found below of just normally media. of blind blind end) end the cotton well a large above University that where cotton better was crossing medium type of conditions right wrapped the around medium. of perithecia of Malaya, CI 3 x 3/8 in the crossing the mouth of ffiola In certain been Lumpur, tend conditions useful. the water. yeast The extract, I% sucrose plus for information Malaya. in I50 for agor cotton (with most ml the in found depending after on the 6 to 8 days regions in the submerged cotton was flasks, the formation the crossing of other Erlenmeyer of slants, fruited in the medium, 3/4”of of crawa. profuse-regions than partially in described in CI number Neurosporo exist When surface technique medium of crossing. in the thick uncovered below kept used perithecia. luxuriantly about l/2” tube with on extra pad of the cotton The tube acted QS a float and crosses, often test-tube fruiting effective to form contained conventional of the inch the mode aerobic on aerial medium the flask. hove crosses agor) and been perithecio found (without few or no perithecia with to ensure particll submergence WCS placed tube towards amount Neurosporo and non-obrortant the was in is thin liquid producing In order which of the cotton medium of the provided layer observed the utilization 85 ml Crosses cottened Botany, in 50 and ogor. publication. production- non-absorbant agor the as Q preservative, (Merck) “and different used, ) is added in the frozen state. N with CI carbon “minimal with be cited (I ml. only. Prokash, use is prepared is dispensed To prepare and Chl oroform 100 ml. room temperature. total volume is about solution is stored ot and a fair and --Department at the with the amount of the strains of