Silver birch (Betula pendula Roth.) is an ecologically and economically important deciduous tree species in Northern Europe. In vitro culture initiation and shoot rejuvenation from birches in their reproductive phase is difficult and often unsuccessful. Currently, markers to evaluate the degree of juvenility of birch in vitro shoots, which could be used to track the rejuvenation process and to determine factors affecting it, have not been developed. The aims of this study were to assess expression of juvenility related microRNAs and target genes during micropropagation of silver birch, and to investigate factors affecting juvenility of birch genotypes with different in vitro morphogenic ability. Potential precursor sequences and target genes of the microRNAs miR156 and miR172 were determined by alignment to the birch unigene set. Ten potential miR156 and miR172 precursor sequences were identified, and were tested for the ability to form the required stem-loop structure. Based on precursor sequences, primers were designed for real time PCR analysis of precursor miRNA expression. Expression patterns of two miR156 family precursors (miR156_511 and miR156_789) and one miR172 precursor (miR172_1931) and two target genes (BpSPL1 and BpAP2) had the best correlation with juvenility/maturity in the analysed in vitro propagated silver birch samples. Expression patterns of these miRNA precursors and target genes were also investigated in samples cultured under different in vitro conditions. This study provides an initial survey of molecular markers for assessment of phase change in birch in vitro micropropagation.

No abstract

Micropropagation can be used as an alternative method for vegetative propagation of mature birch trees in order to increase effectiveness of breeding. In this study the optimal concentration of synthetic cytokinin 6-benzylaminopurine and naturally occurring cytokinin zeatin were determined for shoot proliferation of birch genotype Bau-40-17. Woody Plant Medium supplemented with 6-benzylaminopurine or zeatin at three different concentrations was used. The results showed that Woody Plant Medium supplemented with 1.0 mg L −1 6-bezylaminopuryne gave the best results for shoot multiplication. In the case of natural cytokinin zeatin, 1.0 mg L -1 with addition of 0.1 mg L -1 kinetin gave the best results, but proliferation rate was significantly lower than with synthetic cytokinin. The presence of cytokinin in the medium affected adventitious root formation and resulted in change of pH, which may have caused a positive effect on shoot multiplication rate.

Silver birch (Betula pendula Roth.) is an ecologically and economically important deciduous tree species in Northern Europe. In vitro culture initiation and shoot rejuvenation from birches in their reproductive phase is difficult and often unsuccessful. Currently, markers to evaluate the degree of juvenility of birch in vitro shoots, which could be used to track the rejuvenation process and to determine factors affecting it, have not been developed. The aims of this study were to assess expression of juvenility related microRNAs and target genes during micropropagation of silver birch, and to investigate factors affecting juvenility of birch genotypes with different in vitro morphogenic ability. Potential precursor sequences and target genes of the microRNAs miR156 and miR172 were determined by alignment to the birch unigene set. Ten potential miR156 and miR172 precursor sequences were identified, and were tested for the ability to form the required stem-loop structure. Based on precursor sequences, primers were designed for real time PCR analysis of precursor miRNA expression. Expression patterns of two miR156 family precursors (miR156_511 and miR156_789) and one miR172 precursor (miR172_1931) and two target genes (BpSPL1 and BpAP2) had the best correlation with juvenility/maturity in in vitro propagated silver birch. Expression patterns of these miRNA precursors and target genes were also investigated in samples cultured under different in vitro conditions. This study provides an initial survey of molecular markers for assessment of phase change in birch in vitro micropropagation.