US 11,965,156 B2
RNA isolation from soluble urine fractions
Heather Sanders, Winchester, CA (US); and Hai-Rong Li, Mission Viejo, CA (US)
Assigned to Quest Diagnostics Investments LLC, Secaucus, NJ (US)
Filed by Quest Diagnostics Investments LLC, Secaucus, NJ (US)
Filed on Dec. 7, 2021, as Appl. No. 17/543,944.
Application 17/543,944 is a continuation of application No. 16/692,355, filed on Nov. 22, 2019, granted, now 11,193,120.
Application 16/692,355 is a continuation of application No. 16/118,864, filed on Aug. 31, 2018, granted, now 10,494,629.
Application 16/118,864 is a continuation of application No. 15/130,039, filed on Apr. 15, 2016, granted, now 10,066,226.
Application 15/130,039 is a continuation of application No. 12/973,747, filed on Dec. 20, 2010, granted, now 9,315,802.
Claims priority of provisional application 61/290,976, filed on Dec. 30, 2009.
Prior Publication US 2022/0090049 A1, Mar. 24, 2022
This patent is subject to a terminal disclaimer.
Int. Cl. C12P 19/34 (2006.01); C12N 15/10 (2006.01); C12Q 1/6886 (2018.01)
CPC C12N 15/1017 (2013.01) [C12Q 1/6886 (2013.01); C12Q 2600/118 (2013.01); C12Q 2600/158 (2013.01)] 9 Claims
 
1. A method for amplifying nucleic acids from a urine sample for detection of RNA expression associated with a cancer, comprising;
a) separating urine sediment from the soluble urine fraction of a urine sample obtained from an individual suspected of having cancer, wherein RNA associated with the cancer is present in the soluble urine fraction,
b) concentrating the soluble urine fraction by ultrafiltration using a filter having a nominal molecular weight limit of between about 3,000 daltons and about 100,000 daltons, to produce a soluble urine concentrate, wherein the volume of the soluble urine concentrate is reduced at least 50% from the original urine volume,
c) isolating RNA from the soluble urine concentrate produced in step b),
d) reverse transcribing the RNA from the soluble urine concentrate produced in step c) to form cDNA; and
e) amplifying the cDNA produced in step d), wherein the cancer-associated RNA is selected from the group consisting of prostate cancer antigen 3 (PCA3), p antigen family, member 4 (PAGE4), solute carrier family 45, member 3 (SLC45A3), kallikrein-related peptidase, member 3 (KLK3), phosphoinositide-3-kinase, alpha polypeptide (PIK3CA), erythroblastic leukemia viral oncogene homolog 2 (ERBB2), hyaluronoglucosaminidase (HYAL1), bladder cancer associated protein (BLCAP), solute carrier family 35, member E3 (SLC35E3), tumor protein p53 (TP53), and suppressor of tumorigenicity 20 (ST20).