Date of Award

2017

Document Type

Open Access Master's Report

Degree Name

Master of Science in Computer Science (MS)

Administrative Home Department

Department of Computer Science

Advisor 1

Hairong Wei

Committee Member 1

Zhenlin Wang

Committee Member 2

Myounghoon Jeon

Abstract

Small RNAs including microRNAs (miRNAs) and short interfering RNAs (siRNAs) are widely present in plants. They are transcribed from non-coding small RNA genes and then play as regulators to modulate the levels of messenger RNAs (mRNAs) of protein-coding genes via sequence pairings. This is because a paired complementary double sequence helix structure can trigger mRNA degradation or interfere with mRNA translation. Short Tandem Target Mimic (STTM) is a recently developed technology that can be used to produce a complementary sequence to a miRNA and destroy it or reduce the expression level of this miRNA via the formation of paired double-strand structure. Research has shown success in plant species like Arabidopsis [1] and tomato [2]. The main motivation of this thesis report is to describe a web application named DesignSTTM portal, which has been developed to computerize and automate the design of a STTM sequence that enables one to target a given miRNA species for degradation via sequence complementarities mechanism. The DesignSTTM then inserts the designed STTM sequence immediately downstream of the Cauliflower mosaic virus promoter called 35S in a DNA plasmid vector that can carry 35S promoter + STTM expression cassette into nuclei, and then insert it into a genome, where 35S can drive a STTM sequence to express and produce complementary sequences to target miRNA for silencing. Before the DesignSTTM portal displays the designed plasmid vector carrying STTM sequence in the form of a circular map, it uses an algorithm called basic local alignment search tool (BLAST) to query a locally installed vector database and then annotate all the elements in the plasmid vector. The elements that will be identified and annotated include all the genes, restriction enzyme sites, promoters, and the location of the inserted STTM sequence, which will then be plotted as features of plasmid vector in a map. The portal also displays the plasmid DNA sequence resulting after the insertion of STTM sequence, with a highlight of the location of the STTM sequence. The DesignSTTM portal also has a function to design the primer sequences used to amplify the STTM sequence and majority of plasmid vector using Polymerase Chain Reaction (PCR) technology and transfer them to a binary vector. The DesignSTTM portal is also capable of producing a PDF document incorporating all the derived results, and sending the file as an attachment to users’ email. Blossom STTM Hub’s also maintains an account for each user to store all the obtained results. The other web portal that was implemented in this Blossom STTM Hub’s is called MaterialSTTM, which was designed to store and visualize both genotypic and phenotypic data collected from STTM transgenic lines generated through transformation of STTM plasmid vectors into different plant species. Users can purchase the transgenic seeds available on MaterialSTTM portal by placing an order through the e-commerce store set up through the MTU Touchnet service. This tool will be instrumental for numerous plant biologists who study the functions of miRNAs species through modulating their expression levels in plants.

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