Document Type

Thesis

Date of Award

5-2024

School/College

College of Science, Engineering, and Technology (COSET)

Degree Name

MS in Biology

Committee Chairperson

Audrey Player

Committee Member 1

Erica Cassimere

Committee Member 2

Mario Hollomon

Committee Member 3

Jahmario Williams

Keywords

MultAlin, MYBL1 Gene, PCR, RNA

Abstract

Several years ago our laboratory identified the MYBL1 gene as over-expressed in a subset of triple negative breast cancer cells. Since then we have applied a number of different approaches in an attempt to better define the presence of the gene in these cancers. There is clear and convincing evidence both by other investigators and from experimental observations in our laboratory that alternate splice sequences of a gene can code for variants with different biological roles. We are considering such a duality for the MYBL1 gene in breast samples. The first step towards these studies requires careful analyses of the sequences related to the MYBL1 transcript variants. Data deposited at the National Center for Biotechnology Information show that there are 17 Reference Sequence transcript variants corresponding to the MYBL1 gene. Several of the variants are sequenced as duplicates resulting in 10 unique variants. The goal of our laboratory is to better understand the MYBL1 gene. The current project involves (a) comparative analyses of the sequences assigned to the MYBL1 transcript variants so that (b) primer sets could be designed that would allow for gene expression analyses of the variants in triple negative breast cell line preparations. Both curated and predicted Reference sequences were examined in this study. As expected, the variants demonstrate considerable sequence similarity with the most significant differences observed between the curated transcript variant 1 (NM_001080416.4) which is the longest, and the predicted transcript variant 7 (XM_017013459.2) which is the shortest Reference sequence. Individual exon sequences were also compared between the different transcript variants. The longest variant 1, NM_001080416.4 (along with 3 of the predicted variants) contain an extra exon 15 missing in the other variants. The exon 15 region corresponds to a regulatory region in the protein. We designed 4 sets of PCR primers that allowed for differential analyses between the 10 transcript variants. The primer sets were used to examine the differential MYBL1 transcript levels in a non-tumor triple negative sample, compared to triple negative breast cancer levels. The most significant differential expression between non-tumor and tumor was observed for primer sets that did not detect XM_017013459.2 transcript variant 7. These data suggest that XM_017013459.2 does not contribute to detection of MYBL1 transcripts in tumor cells.

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