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MicroRNA profiling in different types,grades and stages of breast cancer

ABSTRACT: MicroRNA profiling was done using Taqman Low Density Arrays (TLDA) platform consisting of 667 microRNAs covering Sanger miRBase version10 cross 50 samples along with their adjacent normals consisting of different types (ER+ and ER-ve), grades (grade 2 and grade 3) with their different stages (I to III). The above isolated RNA which displayed good RIN value and linearity (R2>0.96), were used for reverse transcription (RT) reactions with the help of TaqMan MicroRNA Reverse Transcription Kit followed by real time PCR Reactions (ABI 7900 HT) as per manufacturer's instructions.A pre -amplification step of cDNA with pre-amp megaplex pool primers was carried out to significantly enhance the ability to detect highly down regulated miRNAs. The TaqMan human microRNA arrays consists of two plates, pool A and pool B. RNU 46 and RNU 48 were used as endogenous controls for data normalization. Another control not related to human was also included as a negative control. Each TaqMan Assay was run in quadruplicate. RNU 46 and RNU 48 expression was consistent in all the samples and displayed good range of CT values (22–24 ) where as in ‘No Template’ Control (NTC) CT value was above 38. The average CT values of total profiled miRNAs in all samples were normalized with RNU 46 & RNU 48 using spotfire (statminer) software and fold changes were represented in terms of 2 - Δ Δ CT (RQ ) and log10RQ. Only valid and significant miRNAs were picked up for further analysis.

ORGANISM(S): Homo sapiens

PROVIDER: GSE144463 | GEO | 2020/01/30

REPOSITORIES: GEO

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MicroRNA profiling in different types,grades and stages of breast cancer

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