cDNA library preparation from a single wheat kernel

• Autorzy: Takacs I. , Koszegi D. , Barnabas B.
• Języki publikacji: EN

Abstrakty

EN

A simplified cDNA cloning protocol was elaborated, easily scalable to very different sizes of plant tissue. The mRNA fraction was extracted from a single wheat kernel with oligo dT magnetic beads. The solid-phase mRNA was transcribed into single-strand cDNA by reverse transcription. Then DNA tags were incorporated into the ds cDNA fragments prior to PCR amplification. The amplified DNA was ligated to a T overhang cloning vector and some of the resulting clones were sequenced. These sequences were identified by BLASTN search in wheat EST databases.

Słowa kluczowe

EN

wheat; kernel; cDNA library; polymerase chain reaction; DNA sequencing; plant tissue; mRNA fraction; BLASTn database; EST database

• Wydawca: -
• Czasopismo: Acta Biologica Cracoviensia. Series Botanica
• Rocznik: 2008
• Tom: 50
• Numer: 1
• Opis fizyczny: p.105-109,fig.,ref.

Twórcy

autor

Takacs I.

• Agricultural Research Instituteof the Hungarian Academy of Sciences, H-2462, Martonvasar, POB.19, Hungary

autor

Koszegi D.

autor

Barnabas B.

Bibliografia

• Aslanidis C, and de Jong P. 1990. Ligation-independent cloning of PCR products. Nucleic Acids Research 18: 6069-6072.
• Clarke BC, Laroque OR, Bekes F, Somers D, and Appels R. 2002. The frequent classes of expressed genes in wheat endosperm tissue as possible sources of genetic markers. Australian Journal of Botany 53: 1181-1193.
• Dresselhaus T, Lörz H, and Kranz E. 1994. Representative cDNA libraries from few plant cells. The Plant Journal 5: 605-610.
• Ikeda TM, Nagamine T, Fukuoka H, and Yano H. 2002. Identification of new low-molecular-weight glutenin subunit genes in wheat. Theoretical and Applied Genetics 104: 680-687.
• Karrer EE, Lincoln JE, Hogenhout S, Bennett AB, Bostock RM, Martineau B, Lucas WJ, Gilchrist DG, and Alexander D. 1995. In situ isolation of mRNA from individual plant cells: Creation of cell-specific cDNA libraries. Proceedings of the National Academy of Sciences USA. 92: 3814-3818.
• Lambert KN, and Williamson VM. 1993. cDNA library construction from small amounts of RNA using paramagnetic beads and PCR. Nucleic Acids Research 21: 775-776.
• McCarry JR, and Williams SA. 1994. Construction of cDNA libraries from limiting amounts of material. Current Opinion in Biotechnology 5: 34-39.
• Nagy IJ, Takács I, Tamás L, and Bedõ Z. 2003. Molecular cloning and characterization of low-molecular-weight glutenin sequences from an old Hungarian wheat variety, Bánkúti 1201. Cereal Research Communication 31: 25-31.
• Sardelli AD. 1993. Plateau effect. Understanding PCR limitations. Amplifications: A Forum For PCR Users 9: 1-5.
• Schmidt WM, and Mueller MW. 1996. Controlled ribonucleotide tailing of cDNA ends (CRTC) by terminal deoxynucleotidyl transferase: a new approach in PCR- mediated analysis of mRNA sequences. Nucleic Acids Research 24: 1789-1791.
• Shibata Y, Carninci P, Watahiki A, Shiraki T, Konno H, Muramatsu M, and Hayashizaki Y. 2001. Cloning full-length cap-trapper-selected cDNAs by using the singlestrand linker ligation method. BioTechniques 30: 1250-1254.
• Stamm S, and Brosius J. 1991. Sanchored PCR: PCR with cDNA coupled to a solid phase. Nucleic Acids Research 19: 1350.
• Zierold U, Scholz U, and Schweizer P. 2005. Transcriptome analysis of mlo-mediated resistance in the epidermis of barley. Molecular Plant Pathology 6: 139-151.