A HlXDREl) MlCROSCOl'K'AL OBJECTS 225 



or if id) there is sometimes an interphase and sometimes 

 not in the same plant. Use preferably smears fixed with 

 chrom-acetic-formalin and stained with iron-brazilin, or 

 iron-hsematoxylin in 70 per cent alcohol. (Sakamura, Bell- 

 ing, Rosenberg, Matsuda.) 



16. Amphidiploid Species Hybrids. — These theoreti- 

 cally interesting plants occur rarely in the first filial genera- 

 tion (or later) in difficult species crosses. Some of these 

 are Nicotiana glutinosa by A^. tabacum; Primula floribunda 

 and P. verticillata; Raphanus and Brassica; Fragaria spp,; 

 Triticum and Aegilops; and Solanum spp. If some of these 

 can be remade, or seeds of them obtained, it is instructive 

 to compare the first metaphase of the constant amphi- 

 diploid hybrid with the metaphases of its two parental 

 species. The cells of the Nicotiana spp. and of their con- 

 stant hybrid stain well if smears are fixed in chrom-acetic- 

 formahn and stained for 2 or 3 hours in iron-brazilin, 

 followed by rather rapid extraction in iron alum. (Clausen 

 and Goodspeed, Karpechenko, Tschermak and Bleier, 

 Ichijima, etc.) 



17. Leptotene, Zygotene, Pachytene, and Diplotene 

 Stages of Allium. — An Allium labelled A. triquetrum was 

 used by the writer and was superior to A . cepa; but perhaps 

 other species are also good. Smears are to be made from 

 the buds at the right stage, fixed for 2 to 3 hours in chrom- 

 acetic-formalin, washed 10 minutes in chrom-acetic, run 

 through the alcohols, mordanted for 24 hours, stained in 

 brazilin for 24 hours, well differentiated, and run through 

 alcohol plus cedar oil and cedar oil plus xylol, into cedar 

 oil. Covers should be 0.17 miUimeter thick. The cells 

 are to be squeezed individually with a flexible penknife 

 blade, so that much of the cytoplasm is squeezed out of 

 the cell, leaving the chromosomes. A 100 fluorite objec- 

 tive, with a condenser giving a corrected cone of 1.2, a 12.5 

 compensating eyepiece, a diaphragm on the radiant making 

 it equal to the source-field, and Wratten screen No. 57A, 

 give nearly optimum results with a sufficiently strong, but 

 not too strong, light. Sometimes the preparations show 

 well, for a time, in hyrax. 



