Molecular cloning and expression of voltagedependent Anion Channel 2 (VDAC2) from electrically stimulated chicken for antibody development
We earlier reported proteomic profiles indicating VDAC2 as a potential biomarker for over-stunned zchicken. The current work focused on cloning VDAC2 gene from chicken’s skeletal muscle into Escherichia coli. R A from chicken’s skeletal muscle was isolated and purified to synthesize the cDNA. Aft...
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| Main Authors: | , , |
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| Format: | Conference or Workshop Item |
| Language: | English English |
| Published: |
2012
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| Subjects: | |
| Online Access: | http://irep.iium.edu.my/26146/1/BT-203_Nur_Hidayah_Hassan_Micotribe_2012.pdf http://irep.iium.edu.my/26146/4/BT203.pdf http://irep.iium.edu.my/26146/ http://micotribe2012.unimap.edu.my/ |
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| Summary: | We earlier reported proteomic profiles indicating VDAC2 as a potential biomarker for over-stunned
zchicken. The current work focused on cloning VDAC2 gene from chicken’s skeletal muscle into
Escherichia coli. R A from chicken’s skeletal muscle was isolated and purified to synthesize the
cDNA. After DNA amplification by proofread DNA polymerase, the PCR product was purified and
inserted into pENTR-TEV-D-TOPO entry vector and transformed in One Shot Chemically
Competent E. coli cells. The entry clone then were inserted into pDEST 17 cloning vector using LR
recombination reaction and E.coli DH5α strain as the host. The vector construct was tranformed
into E. coli BL21A1 strain for protein expression. All positive clones were identified by PCR and
restriction enzyme digestion as well validated by DNA sequencing. Recombinant VDAC2 protein
was purified by Ni-NTA spin-column and Western blot performed on the purified recombinant
VDAC2 protein with anti-His followed by AP goat anti-rabbit revealed a 30 kDa protein band and
confirmed the protein expression. |
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