RESUMO
Lipoic acid synthase, which exists primarily in mitochondria, participates in the biosynthesis of intrinsic lipoic acid. The lipoic acid synthase gene in pig is known as LIAS. To further investigate the biological functions of the protein that is encoded by LIAS, we cloned the open read frame of porcine LIAS (GenBank No. JN797612.1) into the expression vector pET-28α(+). The resulting pET-28α(+)-Lias recombinant vector was introduced into the Escherichia coli BL21 (DE3) strain. With induction by isopropyl ß-D-1-thiogalactopyranoside, the recombinant E. coli strain can express the target protein that has a molecular weight of 41.58 kDa, which was confirmed by Western blotting.
Assuntos
Expressão Gênica , Fases de Leitura Aberta , Sulfurtransferases/genética , Animais , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Peso Molecular , Plasmídeos/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Sulfurtransferases/metabolismo , SuínosRESUMO
Interkeukin-8 (IL-8) is an important inflammatory mediator. It is an angiogenic factor associated with inflammation and carcinogenesis. To date, research on IL-8 has been limited to its role as an indicator of inflammation. There has been no systematic research concerning IL-8 expression levels in the mouse mammary gland during pregnancy and lactation. Mouse mammary gland samples were collected on days 1, 6, 12, 18 of pregnancy and of lactation (6 mice per group). The expression levels of IL-8 mRNA were measured by semi-quantitative RT-PCR, with GAPDH as an internal control. IL-8 mRNA was highly expressed on day 1 of pregnancy in the mouse mammary glands (IL-8(IOD)/GAPDH(IOD) = 1.68), and then suddenly declined; it reached 0.74 and 0.71 on days 6 and 12 of pregnancy. On day 18 of pregnancy, it started to increase (IL-8(IOD)/GAPDH(IOD) = 1.02). However, the expression levels of IL-8 mRNA were not significant during pregnancy. During lactation, IL-8 expression level was lower than during pregnancy, but it stabilized at 0.32-0.41 (IL-8(IOD)/GAPDH(IOD)) from day 1 to day 18 of lactation, although the difference was not significant. We suggest that the changes in IL-8 expression level during development is related to its regulatory role in mouse mammary gland immunity.