Four types of polyacrylamide or polydimethyl-acrylamide gels for regioselective (by immobilization at the 3' end) of short oligonucleotides have been designed for use in manufacturing oligonucleotide microchips. Two of these supports contain amino or aldehyde groups in the gel, allowing coupling with oligonucleotides bearing aldehyde or amino groups, respectively, in the presence of a reducing agent. The aldehyde gel support showed a higher immobilization efficiency relative to the amino gel. Of all reducing agents tested, the best results were obtained with a pyridine-borane complex. The other supports are based on an acrylamide gel activated with glutaraldehyde or a hydroxyalkyl-functionalized gel treated with mesyl chloride. The use of dimethylacrylamide instead of acrylamide allows subsequent gel modifications in organic solvents. All the immobilization methods are easy and simple to perform, give high and reproducible yields, allow long durations of storage of the activated support, and provide high stability of attachment and low non-specific binding. Although these gel supports have been developed for preparing oligonucleotide microchips, they may be used for other purposes as well.

A new procedure has been developed for the synthesis of 3'-amino-3'-deoxyribonucleosides of adenine, cytosine and uracil by condensing the trimethylsilylated bases with peracylated 3-azido-3-deoxyribose derivative. The azido group could subsequently be reduced to amino. The 5'-phosphates of these nucleosides have been prepared and the analogues have been tested for their ability to stimulate the ribosome-catalyzed reaction of 3'(2')-O-(N-formylmethionyl) adenosine 5'-phosphate with phenylalanyl-tRNA.

We have found that the 5'-pdGTT molecules at a concentration of lo-' M HK oligomerired in solution in the presence of lo-' M tripeptide -(L-Val),-N H-NH-DNSCFJOOH and the condensation reagents (carbodiimidc and imidazolc). Oligonuclcotidcs not less than I2 bases long were formed in the yield which was over 15%. It is known that in the absence or peptide IO" M mono-or dinucleotides are required. Thus trivalinc can bc considered as OIIC of the simplest enzymes. This oligomsrization seems to be an essential way for the synthesis of long enough oligonuclcotides of the random GC-sequence, which could be used at the earliest steps of evolution.

The mutant T7 RNA polymerase (T7 RNAP), containing two substitutions (Y639F, S641A) was earlier shown to utilize both rNTP and dNTP in a transcription-like reaction. In this report the ability of the enzyme to catalyze DNA primer extension reaction was demonstrated. The efficiency of the reaction essentially depended on the type of the primer sequence, and was significantly higher if the primer coincided with the T7 promoter non-coding sequence. In this case the primer extension reaction proceeded along with de novo RNA synthesis. The length of the product did not exceed 8 nucleotides, indicating that the primer extension reaction proceeds according to the mechanism of the T7 RNAP-catalyzed abortive transcription.z 1998 Federation of European Biochemical Societies.