| Summary: | 碩士 === 國立臺灣大學 === 生化科學研究所 === 82 === Gamma-Crystallin is a ubiquitous lens protein of most
vertebrate eye lenses and the major component in lenses of
fishes and many mammals, especially during embryonic and
neonatal stages. To date it seems to be absent in the birds and
some reptiles. To facilitate the cloning and sequencing ofγ-
crystallin gene,cDNA was constructed from poly(A)+RNA of shark
lenses (Chiloscyllium colax), amplified by the polymerase chain
reaction (PCR), using the 5'- and 3'- coding regions of carp γ-
crystallins(γs,γm1,γm2) as forward and reverse primers. The
PCR products pretreated with Klenow fragment and polynucleotide
kinase was subcloned into pUC18 vector and then transformed
into E. coli strain JM109. Four positive γ-cry clones of shark
were obtained and processed for nucleotide seq- uencing.
Through sequencing by Sanger's termination method, we have
found these cDNAs can be seperated into two classes; one is
mammalian-like γ-cry, which comprises γs-1,γs-2,γm2 (about
65% homology in protein sequence with bovine γB) with low Met
content (5-6%).The other class is teleostean γ-cry ( about 80%
homology in protein sequence with carp γm1),that isγm1,
unlike γs-1,γs-2,and γm2, which contains high Met con-
tent(-16%). The structural analysis of the crystallins deduces
from these cDNAs is carried out in order to shed some light on
the molecu- lar evolution of these structurally homologous
proteins.Pre- liminary study on the genomic structure encoding
these crystall ins were also presented.
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