Crude cytoplasmic lysate

Incubate 50 fmol [32P]RNAR and 15 xg protein from a crude cytoplasmic lysate in 10 pi binding buffer at room temperature for 15 min. 

A cytoplasmic extract consists of a crude egg lysate containing cytosol, membrane vesicles (MVs), ribosomes, and occasional mitochondria, but is free of pronuclei, essentially all yolk granules, and other large cytoplasmic inclusions. Unfertilized egg extracts support only partial chromatin decondensation (Cameron and Poccia, 1994), whereas extracts from fertilized eggs support full chromatin decondensation and nuclear envelope assembly (Zhang and Ruderman, 1993 Cameron and Poccia, 1994 Collas and Poccia, 1995a,b). 

Figure 2. PemB cellular localisation. (A) Fractionation of E. chrysanthemi cells by spheroplasting. Lane 1, culture supernatant lane 2, total cell lysate lane 3, periplasmic fraction lane 4, crude membrane fraction lane 5, cytoplasmic fraction. (B) Detergent extraction of PemB from E. chrysanthemi A837 cell envelopes. Lane 1 crude envelope fraction lane 2 Triton-soluble fraction lane 3 Triton-insoluble fraction lane 4 Sarkosyl-soluble fraction lane 5 Sarkosyl-insoluble fraction.

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