Isolation of Adaptive Genes from Shorea curtisii Dyer ex King via PCR
Two pairs of primers for heat shock protein 90 (HSP90) and abscisic acid induced guard cell protein (ABGI) which are adaptive genes responsible for heat stress and drought stress, respectively were designed and synthesized based on the sequence of HSP90 in Catharanthus roseus and ABGI in Vicia f...
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| Main Author: | |
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| Format: | Final Year Project Report |
| Language: | English |
| Published: |
Universiti Malaysia Sarawak, (UNIMAS)
2005
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| Subjects: | |
| Online Access: | http://ir.unimas.my/id/eprint/46710/1/Ang%20Swee%20Kim%20ft.pdf http://ir.unimas.my/id/eprint/46710/ |
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| Summary: | Two pairs of primers for heat shock protein 90 (HSP90) and abscisic acid induced guard cell protein
(ABGI) which are adaptive genes responsible for heat stress and drought stress, respectively were designed
and synthesized based on the sequence of HSP90 in Catharanthus roseus and ABGI in Vicia faba. These
primers were used in amplification of HSP90 and ABGI from the genomic DNA of Shorea curtisii via
polymerase chain reaction (PCR). Optimization of PCR produced single distinct DNA fragment with
approximately 750bp HSP90 and 830bp ABGI. Verification of PCR inserts from recombinant plasmids
was conducted via restriction digestion method using EcoR1 and PCR method using the same primer sets,
and also T7/SP6 Promoter primers for further confirmation purpose. Restriction digestion successfully
detected HSP90 PCR insert but failed to detect ABGI PCR insert with desired size. PCR fragments with
size about 750bp and 830bp, which matched the size of HSP90 PCR insert and ABGI PCR insert
respectively, were re-amplified from respective recombinant plasmids using the same primer sets. By using
T7/SP6 Promoter Primers in HSP90 PCR insert and ABGI PCR insert verification, the amplified DNA
fragment was about 920bp and 1000bp respectively. The recombinant plasmids were sent for DNA
sequencing of PCR products in HSP90 and ABGI amplification. The DNA sequencing was in progress and
sequence analysis was not able to be performed. |
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