METHOD OF QUANTITATIVE EVALUATION OF TERMINAL NUCLEOTIDES OF G-STRAND OF TELOMERIC HUMAN DNA BY MEANS OF POLYMERASE CHAIN REACTION AND DUPLEX-SPECIFIC ANALYSIS, SETS OF SYNTHETIC OLIGONUCLEOTIDE PRIMERS AND PROBES FOR IMPLEMENTATION OF ABOVE SAID METHOD
Chiryasova Elena Andreyevna (RU),Чирясова Елена Андреевна (RU),CHIRYASOVA ELENA ANDREYEVNA,Чирясова Елена Андреевна
申请号：
RU2012143263/10
公开号：
RU0002508407C9
申请日:
2012.10.10
申请国别(地区):
RU
年份:
2014
代理人:
摘要：
FIELD: biotechnologies.SUBSTANCE: invention proposes a method involving stages of preliminary extraction of genomic DNA, extraction of a specific fraction of single-stranded G-overhanges of telomeric DNA and further amplification of their negative strand with duplex-specific analysis； besides, amplification stages include modification of 3'-ends of telomeric overhangs by means of terminal deoxynucleotidyl transferase and are implemented using a set of primers SEQ ID NO: 1-5. The duplex-specific analysis is based on hybridisation of minus strands of overhangs with a set of specific fluorescent marked probes SEQ ID NO: 6-11, which correspond to six possible versions of nucleotide endings of G-strand of telomeric human DNA. A buildup degree of probes in presence of a ferment of duplex-specific nuclease serves as a criterion of presence of the corresponding version of the nucleotide ending and a measure of its quantitative content. Obtained profiles of terminal nucleotides of G-strand of telomeric DNA serve as proliferative markers pointing at an activation degree of cell fission and determining time duration of a cellular cycle.EFFECT: invention is a new kind of DNA diagnostics, intended for detection of changes of nucleotide endings of DNA of telomeric areas of chromosomes, which occur in the cellular cycle process, and can be used in medicine for diagnostics and treatment of cancer, in immunology, transplantology, cosmetology, dermatology, gerontology, cellular biotechnology and other areas involved in evaluation of a proliferative state of a cell and methods of its directed control.15 cl, 9 dwg, 8 tbl, 5 exИзобретение относится к молекулярной биологии и генетике клетки. Предложен способ, включающий этапы предварительной экстракции геномной ДНК, выделения специфической фракции однонитевых G-оверхенгов теломерной ДНК и последующей амплификации их минусовой цепи с дуплекс-специфическим анализом, причем этапы амплификации включают модификацию 3'-концов теломерных ове
