Date of Award

1998

Degree Name

Biological Sciences

College

College of Science

Type of Degree

M.S.

Document Type

Thesis

First Advisor

Robert Deal

Second Advisor

Marcia Harrison

Third Advisor

Mary Beth Adams

Fourth Advisor

Leonard J. Deutsch

Abstract

In this study, isoelectric focusing is used to separate proteins of Rhinichthys cataractae and R. atratulus and conventional staining is used to identify alleles of six genes. The objective of this work is to determine whether these methods can effectively assay isozymes that may be used to establish the relationship between anthropogenic stress and genetic diversity. Alleles for LDH-A, LDH-B, EST-A, EST-B, EST-C, and EST-D loci were identified for two species of fish, R. atratulus and R. cataractae. Both fish species are known to be sensitive to anthropogenic stress and known to be widely distributed in stressed environments. Alleles were identified as the isoelectric points for proteins identified as homopolymers. Isoelectric points were calculated relative to protein standards using the regression equations y= 3.8943X+2.49 and y= -1.7277X+6.07 for 3-9 and 5-8 isofocusing gels, respectively. LDH-A focused in the more basic region of the gel, and the LDH-A homopolymer was identified as darkest and widest band produced in the basic region of muscle tissue. The LDH-B homopolymer focused in the more acidic region of the gel. The results were as follow: pl values for R. atratulus, EST-A were 3.82 for muscle, 3.47 for head, and 3.08 for heart tissues. The pl values for R. atratulus, EST-B were 4.13 for muscle, 4.06 for head, and 3.55 for heart tissues. The pl values for R. atratulus, EST-C were 4.84 for muscle, 4.84 for head, and 4.33 for heart. The pl value for R. atratulus, LDH-A was 5.34. The pl values for R. atratulus, LDH-B alleles were 5.37 and 5.18. The pl values for R. cataractae, EST-A were 4.17 for head and heart. The pl value for R. cataractae, EST-B was 4.41. The pl value for R. cataractae, LDH-A was 5.37. The pl values for two LDH-B alleles in R. cataractae were 5.39 and 5.13. Overall, Isoelectric focusing was found to be an accurate and efficient method for the analysis of isozyme diversity of R. cataractae and R. atratulus.

Subject(s)

Isoelectric focusing.

Isoenzymes – Assaying.

Rhinichthys atratulus – West Virginia.

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