Isolation and amplification of DNA sequence coding for peptides with hypocholesterolemic activity from pea (Pisum sativum) and corn (Zea mays)

Date

2011

Degree

Bachelor of Science in Chemistry

College

College of Arts and Sciences (CAS)

Adviser/Committee Chair

Mark Rickard N. Angelia

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Abstract

A list of bioactive peptides exhibiting hypocholesterolemic activity were obtained from various studies. The presence of some of these bioactive peptides in pea and corn were confirmed using In silico analysis. In particular, the peptides IAVP and YVAE were found in the storage proteins of both plants. A forward primer (5'-

CGCAGACCTTACTACTCC-3') and reverse primer (5'-ACTCTTGCTCGTGGTTCC- 3') were synthesized to amplify the target DNA sequence corresponding to the

hypocholesterolemic peptide in the legA gene of pea. A forward primer (5'- CGCAGACCTTACTACTCC-3') and a reverse primer (5'-ACTCTTGCTCGTGGTTCC - 3') were also synthesized to amplify the target DNA sequence in the glb-1 gene of corn. Genomic DNA extraction in pea was done using the method of Cheung et al.- Datta et al.- Dellaporta et al. as modified by Angeles et al. (2005) and the modified Doyle and Doyle protocol was used in the extraction of DNA of corn. The isolated genomic DNA was used as template in PCR. The amplified products from pea had lengths of 550 bp, 400 bp and 290 bp while the amplified product from corn was 800 bp long. Analysis of the PCR products using pairwise alignment showed a 33% homology between the PCR product of pea and the legA gene, while alignment of the PCR product from corn and glb-1 showed 25% similarity. Analysis of the alignment data of the PCR product of pea shows that the product did not correspond to the full length target gene. The result may be attributed to substitution, addition or deletion of bases by the DNA polymerase during amplification. However, a hypocholesterolemic peptide EK was amplified. BLAST search using the PCR product of corn as query showed that it was highly similar to the corn genotype CMS-S mitochondrion. Although the amplified sequence contains the hypocholesterolemic peptide EK, it is still unclear whether the region amplified in the mtDNA can be translated to protein.

Language

English

Location

UPLB Main Library Special Collections Section (USCS)

Document Type

Thesis

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