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What I’ve done this summer Institute of Molecular Biology – Academia Sinica Dr. Che-Kun James Shen 沈哲鯤博士 National Health Research Institutes Dr. Xin Chen 陳新博士 891619 陳惠民 Recent Projects of Dr. Shen Brain cDNA Library Sequencing – Sequencing of macaque’s brain cDNAs – Compare macaque’s brain cDNAs with human’s. – Expect to find some candidate genes which cause the “superiority” of humen over other primates. Brain asymmetrical gene expression – Find some target genes which have notable different gene expression quantity between right and left brains of mice. My work in Dr. Shen’s Lab 20 DNA量 15 左腦X基因 10 右腦X基因 5 0 0 10 20 30 40 PCR cycle數 Learned some concepts of PCR Preliminary screening for target genes derived from right and left brains of mice Qualitative RT-PCR Introductions to Glycophorin Gene Family GPA, GPB and GPE are highly homologous and form a gene cluster on chromosome 4(q28 - 31). The antigens for the MNS blood group system are GPA and GPB. The existence of about 40 variant phenotypes of this blood group system has been documented by serological analyses. Why Are We Interested in Glycophorin Gene Family ? The allelic diversity arises from unequal homologous crossing-over or gene conversions rather than point mutations. The incidence of the allelic diversity across the world appears to be characteristic of the ethnic or geographic origin of the subjects. The evolution of the three identified hot spots. Blood group antigen have become classic genetic markers in genetic population studies and in linkage analyses. Methods PCR with glycophorin-specific primers Blue-white selection Gel electrophoresis Colony PCR Vector-insert ligation Plasmid DNA sequencing Transformation Data analyses Results My work in Dr. Chen’s Lab To learn how to determine the optimal condition for protein purification. Inoculation and incubation overnitht Induction(Different Time, oC and [IPTG]) Ultrasonication centrifucation SDS gel electrophoresis comassive blue staining My work in Dr. Chen’s Lab(Continued) Plasmid construct for future work PCR for the target gene Use restriction enzymes to treat both inserts and vectors. (Check the frame in advance.) Ligation(Check the insert-vector ratio.) Transformation Use the same Restriction enzymes to check whether the insert is inside or not. Plasmid DNA sequencing