title: GSM1705508: Gallus gallus_CHL6_JIIT14DS1_HH40; Gallus gallus; miRNA-Seq
library_construction_protocol: Total RNA was isolated from each six stage of developing chick heart and small RNA was isolated using Ambion mirVana miRNA Isolation Kit. The quality and Integrity of small RNA was checked by Agilent 2000 bioanalyzer. Small RNA cDNA was constructed for each six samples using ScriptSeq RNA-Seq Library Preparation Kit and libraries were constructed . Total six cardiac heart libraries (CHL1 to CHL6) were prepared for deep sequencing and subsequent characterization. For that total RNA samples from each stage were prepared. The low-molecular- weight RNA was precipitated with PEG8000/NaCl. RNA was purified by polyacrylamide gel electrophoresis [PAGE] to elute molecules in the range of 18–30nt. The extracted RNAs were ligated to the 3′ adapter [TCGTATGCCGTCTTCTGCTTG]. The ligated RNAs from each library were eluted from 15% denatured polyacrylamide gel using 0.3N NaCl, followed by the ligation of 5′ adapter [GTTCAGAGTTCTACAGTCCGACGATC]. The samples were further used as templates for cDNA synthesis using small RNA v1.5 cDNA synthesis kit (illumine) by using primers complementary to 5’ and 3’ adapter sequences with 25 PCR cycles of 95°C for 30s, 50°C for 30s, and 72°C for 30s. The PCR products were extracted from the agarose gels using the Gel Extraction Kit [Qiagen]. For each heart developmental stage, equal quantities [20μg] of small RNA were submitted to Illumina Inc.for small RNA deep sequencing.
attributes.tag: GEO Accession
attributes.value: GSM1705508