Biochips – fundamentals and applications - Electrochemical Sensors, Biosensors and Their Biomedical Applications - page 313

TABLE 11.3
Data from inkjet printing method

Dispense volume Spot sizes Spot densities Delivery speed

500-2500 spots/cm 2 50 pL 125-175 µm 100-500 spots/s

droplets and also cause crossover contamination between probes. Due to air bubbles in
inkjet printing, repeatability and reliability of the system may also be greatly reduced.

11.2.3 Sequencing by hybridization
Sequencing by hybridization (SBH) is an approach whereby a collection of overlap-
ping oligonucleotide sequences is assembled together to determine an organism's DNA
sequence. Through the effi cient method of SBH, scientists are able to gather informa-
tion on the genomes of different species and organisms for the future development of
biological sciences, medicine, and agriculture. SBH is based on the complementary
DNA strands' ability to renature after melting as reported by Doty [13]. This results in
the oligonucleotide's probes being hybridized under the condition that allows the com-
plementary sequences in the DNA target to be detected.
In SBH, an m-mer probe is a substring of a DNA sample if it is positively
expressed. This process is similar to doing a keyword search in a page fi lled with text.
The set of positively expressed probes is known as the spectrum of DNA sample. For a
single-strand of DNA sample with the sequence 5 , using a 4-mer probe,

only fi ve probes will hybridize with the single strand DNA. According to Drmanac
et al. [14], all other probes will form hybrids with a mismatch at the end base and
will be denatured during selective washing. The fi ve probes that are of good match at
the end base will result in fully matched hybrids, which will be retained and detected.
Each positively expressed probe serves as a platform to decipher the next base as can
be seen from Fig. 11.6 (see Plate 2 for color version).
In another report by Drmanac et al. [15], Format 1 SBH, a large number of DNA
samples are attached onto nylon membrane, which is used as a solid support to form
the DNA array. These DNA samples can be processed in parallel using a high density
array containing many DNA spots. After repeated cycles of hybridization and washing,
approximately 75% of the initially bonded DNA is retained. The high proportion of
DNA sample retention is due to the formation of covalent interstrand bonds caused by
the UV crosslinking of thymine bases. Following our earlier example of 4-mer probes,
a full set of 256 (or 4 4 ) labeled probes may be scored. Each of these DNA arrays is
exposed to the labeled 4-mer probes and positive hybridizations can be observed at
various DNA spots.
Format 1 SBH can be used to uncover polymorphism and mutations in a particular
gene. The sample amplicons of genomic DNA of a test individual and the amplicons for
the control DNA for the gene of interest with a known reference sequence are both pre-
pared by polymerase chain reaction (PCR). A subset of probes that is complementary

GGTCTCG 3