Detection of microsatellite instability and its use in diagnosis of tumors
View Patent ↗Methods and kits are disclosed for use in the analysis of microsatellite instability in genomic DNA. Methods and kits are also disclosed which can be used to detect microsatellite instability DNA present in biological materials, such as tumors. The methods and kits of the present invention can be used to detect or diagnose diseases associated with microsatellite instability, such as certain types of cancer.
1. A method of detecting microsatellite instability in a DNA sample comprising:
a) co-amplifying a set of microsatellite loci of the DNA sample to produce amplified DNA fragments, the set of loci comprising at least one tetranucleotide repeat locus selected from the group consisting of D1S518, D1S547, D1S1677, D2S1790, D3S2432, D5S2849, D6S1053, D7S3046, D7S1808, D7S3070, D9S2169, D10S1426, D10S2470, D12S391, and D17S1294 wherein the DNA sample is from tumor cells, from cells suspected of being cancerous, or from other biological material suspected of being cancerous;
b) determining the size of the amplified DNA fragments; and
c) comparing the size of the amplified DNA fragments of step b) to the size of corresponding amplified DNA fragments from a paired normal DNA sample, a difference in size between one or more DNA fragments between the samples being indicative of microsatellite instability.
2. The method of claim 1 , wherein the set of microsatellite loci comprises at least two tetranucleotide repeat loci selected from the group consisting of D1S518, D1S547, D1S1677, D2S1790, D3S2432, D5S2849, D6S1053, D7S3046, D7S1080, D7S3070, D9S2169, D10S1426, D10S2470, D12S391, and D17S1294.
3. The method of claim 1 , wherein the set of microsatellite loci comprises at least three tetranucleotide repeat loci selected from the group consisting of D1S518, D1S547, D1S1677, D2S1790, D3S2432, D5S2849, D6S1053, D7S3046, D7S1808, D7S3070, D9S2169, D10S1426, D10S2470, D12S391, and D17S1294.
4. The method of claim 1 , wherein the set of microsatellite loci comprises D1S518, D3S2432, D7S1808, D7S3046, D7S3070 and D10S1426.
5. The method of claim 1 , wherein the microsatellite loci further comprises at least one mononucleotide repeat locus.
6. The method of claim 1 , wherein the set of microsatellite loci is a set of at least five micro satellite loci, comprising:
at least two mononucleotide repeat loci selected from the group consisting of BAT-25, BAT-26, MONO-11, and MONO-15; and
at least three tetranucleotide repeat loci selected from the group consisting of FGA, D1S518, D1S547, D1S1677, D2S1790, D3S2432, D5S818, D5S2849, D6S1053, D7S3046, D7S1808, D7S3070, D8S1179, D9S2169, D10S1426, D10S2470, D12S391, D17S1294, D17S1299, and D18S551.
7. The method of claim 4 , wherein the tetranucleotide repeat loci are amplified using oligonucleotide primers comprising SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:17 and SEQ ID NO:59.