Cloning of the Escherichia coli dapD gene via fragment amplification
| Main Author: | Perpustakaan UGM, i-lib |
|---|---|
| Format: | Article NonPeerReviewed |
| Terbitan: |
[Yogyakarta] : Universitas Gadjah Mada
, 1996
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| Subjects: | |
| Online Access: |
https://repository.ugm.ac.id/17530/ http://i-lib.ugm.ac.id/jurnal/download.php?dataId=289 |
Daftar Isi:
- In Escherichia coli, there are 9 genes responsible for L-lysine biosynthesis from aspartate. One of them is the dapD gene encoding N-succynil diaminopimelate aminotransferase. This enzyme is not found in birds so they are unable to synthesize lysine naturally. To study transformation of the dapD gene into birds, therefore the gene has to be cloned. The main problem of genomic library approach in cloning genes is time consuming. On the other hand, there is a direct cloning approach using amplification fragment by the Polymerase Chain Reaction (PCR) method if the nucleotide sequence of the gene has been known. Base on available data of the E.coli dapD gene sequence, the flanking primers have been made with the addition o/BamHI and EcoRI sites, respectively. The fragment was then amplified by the PCR method. The PCR product was digested with BamHI-EcoRI and then ligated into BamHI-EcoRI digested pUCW. Six out of eight clones analyzed were able to convert E.coli AT982 dapD- to growth without diaminopimelate (DAP). It can be concludeddhat the six clones carry the dapD gene.