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Storage of Bovine Reproductive Tissues and RNA Extracts on Ice for 24 h or Repeated Freeze–Thaw Cycles do not Affect RNA Integrity
Author(s) -
França MR,
Mesquita FS,
Binelli M
Publication year - 2014
Publication title -
reproduction in domestic animals
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.546
H-Index - 66
eISSN - 1439-0531
pISSN - 0936-6768
DOI - 10.1111/rda.12257
Subject(s) - rna , rna extraction , biology , microbiology and biotechnology , messenger rna , gene expression , andrology , chemistry , biochemistry , gene , medicine
Contents The aims of this study were to test (i) the effect of time of tissue and RNA extracts storage on ice and (ii) the effect of repeated freeze–thaw cycles on RNA integrity and gene expression of bovine reproductive tissues. Fragments of endometrium ( ENDO ), corpus luteum ( CL ) and ampulla ( AMP ) were subdivided and incubated for 0, 1, 3, 6, 12 or 24 h on ice. RNA extracts were incubated on ice for 0, 3, 12 or 24 h, or exposed to 1, 2, 4 or 6 freeze–thaw cycles. RNA integrity number ( RIN ) was estimated. Expression of progesterone receptor ( PGR ) and cyclophilin genes from RNA extracts stored on ice for 0 or 24 h, and 1 or 6 freeze–thaw cycles was measured by q PCR . Tissue and RNA extract incubation on ice, and repeated freeze–thaw cycles did not affect RIN values of RNA from ENDO , CL or AMP . Storage on ice or exposure to freeze–thaw cycles did not affect C q values for PGR or cyclophilin genes . In conclusion, neither generalized RNA degradation nor specific RNA degradation was affected by storage of tissue or RNA extracts on ice for up to 24 h, or by up to 6 freeze–thaw cycles of RNA extracts obtained from bovine ENDO , CL and AMP .