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Invitro translation of cytoskeletal beaded-chain filament proteins from chicken lens mRNA

1989; Elsevier BV; Volume: 163; Issue: 1 Linguagem: Inglês

10.1016/0006-291x(89)92178-5

1090-2104

Prabha L. Atreya, H. Maisel,

Molecular Biology Techniques and Applications

The beaded-chain filament is a unique cytoskeletal structure that appears in the elongating fiber cells during the differentiation of lens epithelial cells to form the mature fiber cells. This beaded-chain structure is made up of two proteins of molecular weight 95 kDa and 49 kDa. As a prerequisite for cloning the cDNAs of these proteins, newborn chicken lens total poly(A+) mRNA was translated invitro, using a rabbit reticulocyte lysate system and [35S]-L-methionine. The labelled translation products were analyzed by one-and two dimensional gel electrophoresis followed by autoradiography. Immunoprobing of the translation products on Western blots using specific polyclonal antibodies identified the above proteins, and demonstrated the presence and expression of specific mRNAs in the neonatal chick lens, that code for the invitro synthesis of these two cytoskeletal proteins. These mRNAs are low abundant mRNAs as compared to the crystallin mRNAs.