Production of recombinant protein and polyclonal mouse antiserum for ferritin from Sipuncula Phascolosoma esculenta.
The iron storage protein, ferritin, plays a key role in iron metabolism, but its regulation and functions in many invertebrate species are still largely unknown. In our previous work, an inducible ferritin cDNA from Phascolosoma esculenta with a full-length of 1017 bp has been cloned. In this follow-up study, the deducted ferritin protein sequence was predicted to be a polypeptide of 175 amino acids with a molecular mass of 20.1955kDa and an isoelectric point of 5.08. The cDNA sequence of P. esculenta ferritin was constructed into pET system expression system and efficiently expressed in E. coli BL21 under IPTG induction. The recombinant ferritin was detected as a 24 kDa protein by SDS-PAGE. After purification directly from the gel, the recombinant ferritin was used to immunize mice and the anti-serum was prepared. The antibody displayed a strong immunological reactivity and specificity when used in Western-blot analysis. For the first time, our work provided a set of molecular tools essential for the further studies of ferritin protein functions in P. esculenta.
Su
Xiurong
X
Faculty of Life Science and Biotechnology, Ningbo University, Ningbo, Zhejiang Province 315211, PR China. suxiurong@nbu.edu.cn
Du
Lili
L
Li
Yanyan
Y
Li
Taiwu
T
Li
Dengfeng
D
Wang
Mengqian
M
He
Jingjing
J
D000818:Animals; D000918:Antibody Specificity; D015153:Blotting, Western; D004926:Escherichia coli / Q000235:genetics; D005293:Ferritins / Q000235:genetics / Q000378:metabolism; D007106:Immune Sera; D051379:Mice; D009348:Nematoda / Q000502:physiology*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D007106:Immune Sera; D011994:Recombinant Proteins; D005293:Ferritins