Postdoc 2001 - 2003
I got my Ph.D degree from China
Agricultural University. My previous experiences were mainly trained
on plant molecular genetics, especially molecular markers, such
as RAPD, RFLP, AFLP and microsatellite markers. I was also trained
for rice genomic physical mapping by screening BAC library.
In the Edwards Lab I focused on
evolutionary genomics, with house finches as experimental materials.
The house finch is a common backyard bird throughout most of the
contiguous United States and southeast Canada. Throughout its range,
its fondness for feeding stations and for nesting conspicuously
around buildings make the house finch among North America's most
familiar birds. Several distinctive populations exist within the
native range of the house finch. However, the evolutionary relationships
among these groups needs further study.
In house finches, the disease caused by Mycoplasm
gallisepticum (MG) is manifest as an respiratory and eye infection(conjunctivitis).
House finches naturally infected with MG and held in captivity with
ad libitum food developed conjunctivitis within 2-4 weeks, lost
weight rapidly, and many or all died. From its initial appearance
in the Middle Atlantic States, MG has spread through the entire
population of house finches in the eastern U.S. and southeastern
Canada. The amplified fragment length polymorphism(AFLP) technique
is based on the double amplification via PCR of a subset of restriction
fragments from a total digest of genomic DNA. DNA is cut with two
enzymes (a rare and a frequent cutter according to their restriction
AFLP is a very reproducible technique with a high multiplex ratio,
meaning that a large number of markers can be generated in a single
reaction. By using different combinations of selective primers, an
almost unlimited number of markers can be obtained. The high multiplex
ratio and the fact that no sequence information is needed represent
big advantages over other molecular marker methods.
We employed the AFLP approach to compare
genetic profiles of pre- and post-epidemic finch populations, and
analysis the genetic diversity of house finches on North American
Subtractive hybridization is a powerful
technique, which compare two populations of mRNA and helps obtain
clones that are expressed in one population but not in the other.
Although traditional subtractive hybridization methods have been
successful in some case, they require several rounds of hybridization
and are not well suited to the identification of rare messages.
A new PCR-based cDNA subtraction method, termed suppression subtractive
hybridization (SSH), was first developed by Siebert and colleagues.
SSH is used to selectively amplify target cDNA fragments (differentially
expressed) and simultaneously suppress no target DNA amplification.
The method is based on the suppression PCR effect: long inverted
terminal repeats when attached to DNA fragments can selectively
suppress amplification of undesirable sequence in PCR procedures.
SSH overcomes the problem of differences in mRNA abundance by incorporating
a hybridization step that normalizes (equalizes) sequence abundance
during the course of subtraction by standard hybridization kinetics.
It eliminates any intermediate steps for physical separation of
ss and ds cDNAs, require only one subtractive hybridization round,
and can achieve greater than 1,000-enrichment for differentially
expressed cDNAs. SSH is a highly effective method for generating
subtracted cDNA libraries. It dramatically increases the probability
of obtaining low-abundance differentially expressed cDNAs and simplifies
the analysis of the subtracted cDNA libraries.
Microarrays containing cDNA clones
have been used to compare patterns of gene expression in which thousands
of genes can be examined in a single hybridization. The emerging
technology of cDNA microarry hybridization offers the possibility
for providing a rapid, high throughout method to screen an SSH cDNA
We will incorporate SSH, cDNA microarry
and cDNA-AFLP approaches to identify candidate genes
or markers related to MG disease in house finches.
University of Washington
Seattle, WA 98195