Comparison of complex DNA mixtures with generic oligonucleotide microchips.
The reproducibility of melting curves for repeated hybridizations of target DNA with generic oligonucleotide microchips is shown experimentally to depend on the character of matching between fragments of target DNA and immobilized oligonucleotides. The reproducibility of melting curves is higher for the perfect match duplexes and decreases as the number of mismatched pairs within duplexes increases. This effect was applied to the comparative analysis of complex DNA mixtures. We developed a scheme in which we can identify and discriminate between the probe oligonucleotides responsible for the distinctions between target DNA mixtures. A scheme is illustrated by comparing DNA mixtures corresponding to V-D-J genes connected with populations of mRNAs CDR3 TCR Vb (T-cell receptor beta complementarity determining region 3) from the thymus and pancreas of NOD mice. Our results demonstrate that generic microchips can be applied efficiently to the analysis of DNA mixtures.
Lebed
J B
JB
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow.
Chechetkin
V R
VR
Turygin
A Y
AY
Shick
V V
VV
Mirzabekov
A D
AD
D000818:Animals; D001483:Base Sequence; D003199:Computer Systems; D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification; D004272:DNA, Mitochondrial / Q000737:chemistry; D005782:Gels; D005803:Genes, Immunoglobulin; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D008856:Microscopy, Fluorescence; D008904:Miniaturization; D008969:Molecular Sequence Data; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry*; D020411:Oligonucleotide Array Sequence Analysis; D010179:Pancreas / Q000737:chemistry; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics; D015203:Reproducibility of Results; D013696:Temperature; D013950:Thymus Gland / Q000737:chemistry
D004272:DNA, Mitochondrial; D005782:Gels; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D016693:Receptors, Antigen, T-Cell, alpha-beta; D004247:DNA