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Use of chromosome walking in discovery of single-nucleotide polymorphism in noncoding regions of a candidate actin gene in Pinus radiata

100%

Li W. , Li H. , Wu H. , Chen X.-Y.

Journal of Applied Genetics

2010

tom 51

nr 3

275-281

Untranslated regions (UTRs) of eukaryotic mRNAs play crucial roles in post-transcriptional regulation of gene expression via the modulation of nucleocytoplasmic mRNA transport, translation efficiency, subcellular localization, and message stability. Single-nucleotide polymorphisms (SNPs) in UTRs of a candidate gene may also change the post-transcriptional regulation of a gene or function by nucleotide mutation. For species that have not been entirely sequenced genomically, new methods need to be devised to discover SNPs in noncoding regions of candidate genes. In this study, based on the expressed sequence tag (EST) of Pinus radiata (Monterey pine), we obtained all the sequences of UTRs of the actin gene by using a chromosome walking method. We also detected all the SNPs in and around the coding region of the actin gene. In this way, the full genomic sequence (2154 bp) of the actin gene was identified, including the 5'UTR, introns, the coding sequence, and the 3'UTR. PCR amplification and DNA fragment sequencing from 200 unrelated P. radiata trees revealed a total of 21 SNPs in the actin gene, of which 3 were located in the 5'UTR, 3 in the introns, 10 in the coding sequence, and 5 in the 3'UTR. We show that chromosome walking can be used for obtaining the sequence of UTRs, and then, based on this sequence, to discover SNPs in the noncoding regions of candidate genes from this species without an entire genomic sequence.

Effect of Low Frequency Magnetic Field on Germination of Onion (Allium cepa L.) Seeds

75%

Kubisz L. , Hołubowicz R. , Gauza M. , Li H. , Hojan-Jezierska D. , Jaroszyk F.

nr 1A

A-49-A-53

Commercial seed lots of onion (Allium cepa L.) of the cultivars "Octavia" and "Eureka" were soaked for 12 h in water of the temperature 20°C, then they were exposed to low frequency magnetic field (20 mT) for 10, 30, and 60 min. The field was generated using a standard device for magnetic stimulation VIOFOR JPS applied in human medical treatment. Next, the seeds were evaluated for energy and capacity of germination, length of the received germs as well as fresh masses of the obtained seedlings using the routine methods recommended by the International Seed Testing Association. For the seeds of the cultivar "Octavia", the field did not affect the energy of germination, however it did improve in all tested treatments their germination capacity. In the best of them (60 min), the germination capacity increased by 4.6%. For the seeds of the cultivar "Eureka", the tested field increased their energy of germination, which improves evenness of plants emergences in the field and has a significant importance for horticultural practice. When the seeds were exposed to the field for 60 min, their germination energy increased from 40% (control) to 63%. This result was accompanied by no differences in both length of germs and fresh masses of the received from them seedlings.