BAC-FISH Based Physical Map of Endangered Catfish Clarias magur for Chromosome Cataloguing and Gene Isolation through Positional Cloning.

Construction of a physical chromosome map of a species is important for positional cloning, targeted marker development, fine mapping of genes, selection of candidate genes for molecular breeding, as well as understanding the genome organization. The genomic libraries in the form of bacterial artificial chromosome (BAC) clones are also a very useful resource for physical mapping and identification and isolation of full-length genes and the related cis acting elements. Some BAC-FISH based studies reported in the past were gene based physical chromosome maps of Clarias magur (magur) to understand the genome organization of the species and to establish the relationships with other species in respect to genes' organization and evolution in the past. In the present study, we generated end sequences of the BAC clones and analyzed those end sequences within the scaffolds of the draft genome of magur to identify and map the genes bioinformatically for each clone. A total of 36 clones mostly possessing genes were identified and used in probe construction and their subsequent hybridization on the metaphase chromosomes of magur. This study successfully mapped all 36 specific clones on 16 chromosome pairs, out of 25 pairs of magur chromosomes. These clones are now recognized as chromosome-specific makers, which are an aid in individual chromosome identification and fine assembly of the genome sequence, and will ultimately help in developing anchored genes' map on the chromosomes of C. magur for understanding their organization, inheritance of important fishery traits and evolution of magur with respect to channel catfish, zebrafish and other species.

Thermal stress induces expression of Nuclear protein and Parkin genes in endangered catfish, Clarias magur.

Fluctuation in water temperature can create thermal stress, which may impact many aspects of fish life, such as survival, growth, reproduction, disease occurrence etc. The endangered catfish, Clarias magur, has been reported to survive at higher thermal stress, even though the exact mechanism is unknown. The genes coding for Nuclear protein 1 (Nupr1) and Parkin E3 ubiquitin protein ligase (Park2) have been reported to protect cells from stress-induced damage and death. In this study, we characterized both the genes and assessed their quantitative expression in C. magur. Structural features of both the genes were found similar to a related catfish, Ictalurus punctatus, and model fish zebrafish. The genes were fairly conserved in fishes as observed through phylogenetic analysis. The real time expression of the two stress-associated genes were also assessed in brain, kidney, liver and muscle tissues of C. magur exposed to warm (34 °C) and cold (15 °C) water. RT-PCR analysis revealed up-regulation in the relative expression levels of Nupr1 and Park2 genes at both temperatures with maximum positive fold change during stress to cold water, even though the posteriori Dunnett's test after ANOVA revealed that there were significant differences between the control and challenged groups. The study indicated that Nupr1 gene plays role in muscle tissue at both high and low thermal stress, but at high thermal stress in liver, while Park2 plays role in muscle, brain and kidney at low temperature and in liver at high temperature stress in C. magur. The study has generated first-hand information under warm- and cold water, which pave the way to understand the expression response of these genes to thermal vacillations and to establish evolutionary significance in catfishes and other species.