130
HERMAN
DENIS
Even after a continuous exposure to
3 2
P, ribosomal RNA is, therefore,
much less radioactive than the other types of RNA. The overall specific
activity of
3 2
P-RNA is thus different from that of complementary RNA
and cannot be used to determine the percentage of DNA that can be
saturated by RNA from a given stage.
The difficulty was overcome with a method developed by Brown and
Littna (1966a). This method consisted in determining the specific activity
of the direct precursors of mRNA, i.e., the nucleoside 5'-monophosphates.
These were isolated and purified from the acid-soluble pool of the
embryos. When the eggs are labeled by injecting
3 2
P into the female, the
concentration of the isotope in the inorganic pool of the embryos remains
relatively constant until stage 30-33. After this stage, a considerable
dilution of the isotope occurs, owing to the digestion of the endodermal
yolk, which releases considerable amounts of nonlabeled phosphate.
During a large part of development (stages 0-33), mRNA is, thus, synthesized from precursors the specific activity of which changes relatively little (Brown and Littna, 1966a). After a
3 2
P labeling, it is,
therefore, safe to assume that the specific activity of mRNA is the
same as that of the free nucleotides.
Brown and Littna's method is valid not only when the embryos are
continuously labeled with
3 2
P, but also when they are subjected to a
short pulse with
1 4
C0 2 , followed by a long chase (10 hours) in normal
medium. After a
1 4
C0 2 pulse, the RNA precursors remain radioactive
for at least 30 hr after the end of the pulse (see Figs. 12 and 13). The
specific activity of the free nucleotides reaches a maximum at the end
of the pulse and decreases slowly (a few per cent per hour) during the
chase (Denis, 1967). During the experiment, mRNA is thus synthesized
from precursors the specific activity of which changes slowly. In view of
the relatively fast turnover rate of mRNA, it can be assumed that
mRNA reaches the same specific activity as its direct precursors at the
end of the labeling period.
Ribonucleic acid extracted from growing embryos exposed to
3 2
P from
the beginning of development saturates an increasing percentage of DNA
(Fig. 10). Messenger RNA present in developing embryos is, thus, complementary to an increasing number of DNA sites.
Embryonic
development
is concomitant
with a progressive release of
the genetic information
enclosed in DNA. Except in gastrulae, RNA
labeled with
1 4
C0 2 for 11 hr saturates a smaller percentage of DNA than
RNA from the same stage continuously labeled with
3 2
P (Fig. 10). This
HERMAN
DENIS
Even after a continuous exposure to
3 2
P, ribosomal RNA is, therefore,
much less radioactive than the other types of RNA. The overall specific
activity of
3 2
P-RNA is thus different from that of complementary RNA
and cannot be used to determine the percentage of DNA that can be
saturated by RNA from a given stage.
The difficulty was overcome with a method developed by Brown and
Littna (1966a). This method consisted in determining the specific activity
of the direct precursors of mRNA, i.e., the nucleoside 5'-monophosphates.
These were isolated and purified from the acid-soluble pool of the
embryos. When the eggs are labeled by injecting
3 2
P into the female, the
concentration of the isotope in the inorganic pool of the embryos remains
relatively constant until stage 30-33. After this stage, a considerable
dilution of the isotope occurs, owing to the digestion of the endodermal
yolk, which releases considerable amounts of nonlabeled phosphate.
During a large part of development (stages 0-33), mRNA is, thus, synthesized from precursors the specific activity of which changes relatively little (Brown and Littna, 1966a). After a
3 2
P labeling, it is,
therefore, safe to assume that the specific activity of mRNA is the
same as that of the free nucleotides.
Brown and Littna's method is valid not only when the embryos are
continuously labeled with
3 2
P, but also when they are subjected to a
short pulse with
1 4
C0 2 , followed by a long chase (10 hours) in normal
medium. After a
1 4
C0 2 pulse, the RNA precursors remain radioactive
for at least 30 hr after the end of the pulse (see Figs. 12 and 13). The
specific activity of the free nucleotides reaches a maximum at the end
of the pulse and decreases slowly (a few per cent per hour) during the
chase (Denis, 1967). During the experiment, mRNA is thus synthesized
from precursors the specific activity of which changes slowly. In view of
the relatively fast turnover rate of mRNA, it can be assumed that
mRNA reaches the same specific activity as its direct precursors at the
end of the labeling period.
Ribonucleic acid extracted from growing embryos exposed to
3 2
P from
the beginning of development saturates an increasing percentage of DNA
(Fig. 10). Messenger RNA present in developing embryos is, thus, complementary to an increasing number of DNA sites.
Embryonic
development
is concomitant
with a progressive release of
the genetic information
enclosed in DNA. Except in gastrulae, RNA
labeled with
1 4
C0 2 for 11 hr saturates a smaller percentage of DNA than
RNA from the same stage continuously labeled with
3 2
P (Fig. 10). This