pyrG of Aspergillus nidulans, meiotic mapping, marker interactions and growth response

Information of the genetic location of pyrG and its growth response under various conditions has become important with the recent cloning of pyrG (Oakley et al. 1987. Gene 61:385-399) and with the use of pyrG strains as recipients for transformation when pyr-4 cloning vectors are used for A. nidulans libraries (e.g. May et al. 1985 J. Cell Biol. 101:712-719 ; Osmani et al. 1987 J. Cell Biol. 104:1495-1504). This regular paper is available in Fungal Genetics Reports: https://newprairiepress.org/fgr/vol35/iss1/6 All these mutants are being deposited at FGSC (Table 2) and additional detailed information will be made available to anyone interested in analyzing them further (their investigation is being discontinued). Table 2. New mus strains available from FGSC mus Prototroph strains Simple requiring strains Level of FGSC no. Marker FGSC no. Gene Allele backcross A a gene allele A a mus-7 FK116 II(A) I(a) 6401 6402 ----mus-9 FK129 IV 6403 6404 leu-3 R156 6405 -nic-2 43002 6407 6408 mus-11 FK117 IV 6409 6410 pan-l 5531 6411 6412 lys-1 33933 6413 -mus-21 FK121 VI 6414 6415 trp-1 10575 6416 6417 FK120 II 6418 6419 trp-1 acr-2 10575 KH5 6420 6421 6422 6423 --6424 6425 nic-3 Y31881 6426 6427 mu-27 FK124 IV 6428 6429 arg-5 27947 6430 6431 nuc-2 T28M2 6432 6433 mus-28 FK118 IV 6434 6435 leu-1 33757 6436 6437 mus-29 FK119 IV 6438 6439 trp-2 41 6440 6441 ylo-1 pan-2 Y30539y Y153M96 6442 6443 mus-30 FK115 IV 6444 6445 pan-l 5531 6446 6447 met-2 K43 6448 6449 mus(FK125) IV 6450 6451 rib-l 51602(t) 6452 6453 pan-l 5531 6454 6455 mus(FK128; IV -6457 lys-5 DS6-85 6458 -mus(FK131) II 6459 6460 trp-2 41 6461 6462 mus(FK132) II 6463 6464 rib-l 51602(t) 6465 6466 mus(FK133) II 6467 6468 met-1 M105 6469 6470 Tests of mus(FK115), (FK119) and (FK123) for allelism to recently mapped genes by Dr. H. Inoue are gratefully acknowledged. This work was supported by NSERC of Canada. Biology Dept., McGill University, 1205 Avenue Docteur Penfield, Montreal, Quebec, H3A lB1, Canada Käfer, E. and G. May pyrG of Aspergillus nidulans, meiotic mapping, marker interInformation of the genetic location of pyrG and its growth response under various conditions has become important with the recent cloning of pyrG (Oakley et al. 1987. Gene 61:385-399) and with the use of pyrG strains as recipients for transformation when pyr-4 cloning vectors are actions and growth response. used for A. nidulans libraries (e.g. May et al. 1985 J. Cell Biol. 101:712-719 ; Osmani et al. 1987 J. Cell Biol. 104:1495-1504). Several problems have surfaced in crosses with pyrG89. Two of them, which are related to the genetic mapping of pyrG and were investigated in detail are the following: 1) pyrG is linked to galD but the distance between these markers and orientation of the linked pair were found to vary in crosses with different outside markers; 2) pyrG mutants interact with the linked distal markers, fpaB and trpB, to give very poorly or non-viable double mutant progeny. Two further unexpected problems, encountered among pyrG progeny from heterozygous crosses, were only partly analysed and preliminary results have led to the following proposals: 3) pyrG89 is apparently cold sensitive (cs) on the simple yeast extractglucose supplemented with uridine which is suitable for growth of pyrG recipient strains (YAGU; Osmani et al. 1987, ref. cit.); however, some stocks (e.g., FGSC A576) carry unlinked suppressors which results in 1:1 segregation for cs among pyrG progeny; 4) Two pyrG89 strains (including the Glasgow strain, G191) when crossed to one of two related galD and/or uvsF strains produced a fraction (>1/4) of progeny with a new requirement which can be satisfied by NH4Cl and partly by adenine, but not by nitrate or nitrite. It seems likely that the two cases are related and that the mutations involved are present in stock strains (information about similar observations would be helpful for the assessment and investigation of this intriguing observation). From our extensive analysis of the first two of these problems we conclude that the originally observed differences in linkage values for pyrG were caused by environmental variation rather than chromosomal aberrations. This is demonstrated in Table 1 where results from repeats of the same cross, carried out in different laboratories, show very large differences. On the other hand, the results summarized in Table 2 are based on original data from many different crosses which were relatively uniform (hence the small SEM). Furthermore, when crosses are grouped according to branches of the pedigrees which might involve presumptive normal vs. potential aberration strains, no significant differences in recombination frequencies are seen. The difference in orientation may also partly be caused by variations in conditions (media, crowding, temperature), since the variably poor recovery of the recombinant pyrG fpaB types reduces only one of the two potential double crossover categories; this can create a sufficient bias to apparently reverse the order of pyrG and galD (see A, Table 1). However, the main problem in establishing the orientation of this pair of markers is not specific to pyrG and results directly from the absence of interference in Aspergillus crosses (Käfer 1977 Adv. Genet. 19:33-131).

Käfer, E. and G. May pyrG of Aspergillus nidulans, meiotic mapping, marker inter-Information of the genetic location of pyrG and its growth response under various conditions has become important with the recent cloning of pyrG (Oakley et al. 1987. Gene 61:385-399) and with the use of pyrG strains as recipients for transformation when pyr-4 cloning vectors are actions and growth response.
used for A. nidulans libraries (e.g. May et al. 1985 J. Cell Biol. 101:712-719 ;Osmani et al. 1987J. Cell Biol. 104:1495-1504. Several problems have surfaced in crosses with pyrG89. Two of them, which are related to the genetic mapping of pyrG and were investigated in detail are the following: 1) pyrG is linked to galD but the distance between these markers and orientation of the linked pair were found to vary in crosses with different outside markers; 2) pyrG mutants interact with the linked distal markers, fpaB and trpB, to give very poorly or non-viable double mutant progeny.
Two further unexpected problems, encountered among pyrG progeny from heterozygous crosses, were only partly analysed and preliminary results have led to the following proposals: 3) pyrG89 is apparently cold sensitive (cs) on the simple yeast extractglucose supplemented with uridine which is suitable for growth of pyrG recipient strains (YAGU;Osmani et al. 1987, ref. cit.); however, some stocks (e.g., FGSC A576) carry unlinked suppressors which results in 1:1 segregation for cs among pyrG progeny; 4) Two pyrG89 strains (including the Glasgow strain, G191) when crossed to one of two related galD and/or uvsF strains produced a fraction (>1/4) of progeny with a new requirement which can be satisfied by NH4Cl and partly by adenine, but not by nitrate or nitrite. It seems likely that the two cases are related and that the mutations involved are present in stock strains (information about similar observations would be helpful for the assessment and investigation of this intriguing observation).
From our extensive analysis of the first two of these problems we conclude that the originally observed differences in linkage values for pyrG were caused by environmental variation rather than chromosomal aberrations. This is demonstrated in Table 1 where results from repeats of the same cross, carried out in different laboratories, show very large differences.
On the other hand, the results summarized in Table 2 are based on original data from many different crosses which were relatively uniform (hence the small SEM). Furthermore, when crosses are grouped according to branches of the pedigrees which might involve presumptive normal vs. potential aberration strains, no significant differences in recombination frequencies are seen.
The difference in orientation may also partly be caused by variations in conditions (media, crowding, temperature), since the variably poor recovery of the recombinant pyrG fpaB types reduces only one of the two potential double crossover categories; this can create a sufficient bias to apparently reverse the order of pyrG and galD (see A, Table  1).
However, the main problem in establishing the orientation of this pair of markers is not specific to pyr G and results directly from the absence of interference in Aspergillus crosses (Käfer 1977 Adv. Genet. 19:33-131 Analysis of the distribution of crossing over in the "standard" crosses (of Table 2) confirms earlier meiotic data and indicates a random coincidence with no hint of positive interference.
[Among 81 confirmed cases of crossing over between galD and pyrG, the following fractions of double crossovers were found for adjacent intervals: 4/54 (7.4% for suA, 6/32 (18.8%) for fpaB, and 15/42 (35.7%) for uvsF which in each case is very close to expectation for random coincidence.] When this is the case, two closely linked markers like galD-pyrG (average 3%, Table 2) can reliably be arranged in sequence only if an outside marker is reasonably close. For example, suAadE, at a distance of less than 10%, mapped closer to pyrG than galD in all crosses.
In contrast, the more distant markers fpaB and uvsF usually but not always showed closer linkage to galD in individual crosses.
In general, therefore, when markers at suitable distances are not available, mapping results from single crosses and samples of limited size must remain provisional until confirmed (or reversed, as occurred for several published cases, e.g. galD which originally was placed proximal to suAadE). Table 2 Frequencies of recombination (average % ± SEM) in groups of closely related crosses heterozygous for pyrG and galD and various outside markers.

Outside markers
No.