Biology Reference
In-Depth Information
steps individually, and this conclusion may be viewed as a corollary of the postulate
that the cell is the smallest DNA-based molecular computer (Ji 1999a), or
the
12.6 The Transcription-Transcript Conflation
The DNA microarray technique can be used to measure either DNA or RNA inside
the cell (Fig.
12.5
). Measuring DNA is relatively simple, since all that needs to be
done in this case is to break the cell membrane, transfer the cellular DNA (see D
C
in
Fig.
12.5
) into a test tube (see Step 9), and hybridize it with the DNA (called the
probe
DNA) covalently attached to the surface of a microarray (see Step 5). But
measuring RNA is much more complex, involving at least eight key steps or
processes (see Steps 1-8 explained in the legend to Fig.
12.5
). In other words, the
signal S measured with microarray from an RNA sample isolated from a cell
population is a function of at least eight parameters as shown in Fig.
12.5
:
S
1½
R
C
(12.2)
¼
K[R
C
(12.3)
The Cell
7
4
5
6
3
D
C
R
C
R
T
D
T
D
M
S
1
2
8
Degradation
9
Fig. 12.5 The key steps involved in measuring intracellular RNA levels (
R
C
) with microarrays.
D
c
¼
DNA inside the cell;
R
c
¼
RNA inside the cell;
R
T
¼
RNA isolated in a test tube;
D
T
¼
DNA reverse transcribed from R
T
inside a test tube;
D
M
DNA hybridized to the probe DNA on
the surface of a microarray;
S ¼
the fluorescence signal measured form D
M
.1
¼
transcription;
2
¼
transcript degradation; 3
¼
isolation of RNA from cell (C) into a test tube (T); 4
¼
DNA
synthesis from RNA catalyzed by reverse transcriptase in test tube; 5
¼
hybridization of D
T
to
DNA
probes
covalently attached to the microarray (M) surface; 6
¼
measuring the fluorescence
signal from the
target
DNA hybridized to the probe DNA on the microarray surface; 7
¼
an RNA
molecule affecting the transcription of its own or other genes, either directly (via microRNA, for
example) or indirectly through protein synthesis; 8
¼
an RNA molecule influencing its own
degradation either directly as RNA or through protein synthesis; 9
¼
measurement of cellular
DNA after isolation into a test tube
¼
