Genetic Differentiation of the Blue Swimming Crab Portunus pelagicus Along the Coastal Thai Waters Revealed by SSCP Analysis of Cytochrome c Oxidase Subunit I

The basic information on genetic diversity and population structure is essential for the construction of appropriate management schemes leading to sustainable fisheries of the blue swimming crab (Portunus pelagicus). Here, genetic heterogeneity of P. pelagicus (N=174) was examined by single-strand conformational polymorphism (SSCP) analysis of mitochondrial cytochrome c oxidase subunit I (PpCOI270). Seven SSCP genotypes were found across all investigated samples. The average genetic distance between pairs of geographic samples was 0.0014-0.7247. Significant geographic heterogeneity (P<0.05) and restricted levels of female gene flow between paired samples (0.03-1.60 individuals per generation) were observed except between Chanthaburi - Prachuap Kriri Khan and Ranong - Krabi (P>0.05; 6.54 and 16.17 individuals per generation) located in the same coastal regions. Therefore, the gene pool of P. pelagicus in Thai waters was genetically differentiated to different stocks even though it is biologically regarded as a potential dispersal species. Five geographic samples of P. pelagicus in Thai waters could be differentiated to three genetic stocks; Chanthaburi and Suratthani (stock A), Prachuap Khiri Khan (stock B) and Ranong and Krabi (stock C).

PCR-SSCP Analysis of Keratin - Associated Protein (KAP) 6.1 Gene in Nilagiri and Dorset x Nilagiri Cross Breeds of Sheep

The present study was investigated to detect polymorphic patterns of the keratin-associated protein (KAP) 6.1 gene in 54 numbers of Nilagiri and 20 numbers of Dorset x Nilagiri breeds of sheep. DNA was isolated and amplified with ovine specific primers of KAP 6.1 gene. Allele frequencies of A, B, C, D and E was 0.79, 0.08, 0.15, 0.07 and 0.08 respectively with departure from Hardy-Weinberg equilibrium. KAP 6.1 gene was found to have a high degree of homozygosity (0.6668), and an effective number of alleles (Ne) for KAP 6.1 gene was 1.7007 in Nilagiri breeds of sheep, while the PIC value was 0.3909 with positive (0.1908) FIS value. From the study, PCR - SSCP analysis of KAP 6.1 gene revealed polymorphism in Nilagiri breeds whereas in Dorset x Nilagiri crossbred sheep two genotypes for the A/B with no polymorphism detected.

Variation in lactoferrin gene affects milk lactoferrin content and somatic cell count in murrah buffaloes

Lactoferrin plays an important role in antimicrobial defence and is a potential candidate gene for mastitis resistance. In the present investigation, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) studies were carried out in Murrah (Bubalus bubalis) buffaloes to detect single nucleotide polymorphisms (SNPs) of lactoferrin gene and to analyse the association between the observed polymorphisms with milk lactoferrin content and Somatic cell count (SCC). PCR-SSCP analysis revealed a total of 07 different variants in the partial coding region of the lactoferrin gene. PCR-SSCP analysis of exon 10 of lactoferrin gene revealed three and that of exons 4 and 5 revealed two unique patterns each, while all other exons exhibited monomorphic pattern. Comparison of nucleotide sequences of lactoferrin gene of the Murrah buffaloes with taurine reference sequence revealed a total of 14 point mutations, 09 of which were found to be in coding region. Conceptualized translation of nucleotide sequence revealed 07 amino acid changes. SSCP variants of exon 10 (P less than 0.01) had significant effect on milk SCC. The SSCP variants of exon 4 and exon 5 had significant (P less than 0.05) effect on lactoferrin content. The SCC and lactoferrin content in Murrah buffaloes was highest in 4th and above parity group. Stage of lactation had highly significant (P less than 0.01) effect on both milk SCC and lactoferrin content. There was a high and significant (p less than 0.01) correlation (0.741) between SCC and lactoferrin content in milk. The observed association between SSCP variants in lactoferrin gene with milk SCC and milk lactoferrin content can be used as prognostic markers for selection of animals for high lactoferrin content and low somatic cell count, as well as a marker of susceptibility/resistance to mastitis in Murrah buffaloes

“Mirror” Method to Estimate Mutagenic Activity of DNA Lesions

We propose an improved method for detecting mutations that arise in DNA due to misincorporations of deoxyadenosine-5′-monophosphate (dAMP) opposite 7,8-dihydro-8-oxoguanine (8-oxoG). The method is based on the synthesis of complementary chains (“mirror” products) using a template containing 8-oxoG. The misincorporation of dAMP in the “mirror” product forms EcoRI sites. The restriction analysis of double-stranded DNAs obtained by PCR of “mirror” product allows quantification of the mutagenesis frequency. In addition, single-strand conformational polymorphism (SSCP) analysis of the single-stranded “mirror” products shows that different DNA polymerases only incorporate dA or dC opposite 8-oxoG. The proposed approach used in developing this technique can be applied in the study of other lesions as well, both single and clustered.