Total RNA was extracted from sorted bone marrow derived macrophages, lung AM and brain microglia of WT or Csf1rCreDicerfl/fl knockout mice with miRNeasy Mini Kit (QIAGEN). The RNA was reverse-transcribed to cDNA with miRCURY LNATM microRNA RT Kit (Exiqon). Quantitative real-time PCR reactions were prepared using FastStart Universal SYBR Green Master (ROX, Roche) and carried out in QuantStudio 7 Flex Real-Time PCR System (Applied Biosystems). The U6 primer set was purchased from Exiqon (Product No.: 203907). All other miRNA primer sets were purchased from Life Technologies, including miR-17 (Assay ID: 002308), miR-18a (Assay ID: 002422), miR-19a (Assay ID: 000395), miR-19b (Assay ID: 000396), miR-20a (Assay ID: 000580), miR-92a (Assay ID: 000430), miR-21 (Assay ID: 000397), miR-146a (Assay ID: 000468), miR-150 (Assay ID: 000473), miR-155 (Assay ID: 002571) and miR-233 (Assay ID: 002295). Relative miRNA expressions were normalized to U6 expression.
Mir 19b
Manufactured by Thermo Fisher Scientific
Sourced in Spain, United States
MiR-19b is a microRNA (miRNA) product from Thermo Fisher Scientific. miRNAs are small, non-coding RNA molecules that play a role in regulating gene expression. This product provides the miR-19b miRNA sequence.
Automatically generated - may contain errors
Lab products found in correlation
Mir 17, by Thermo Fisher Scientific (1 mentions)
Mir 21, by Thermo Fisher Scientific (1 mentions)
Mir 146a, by Thermo Fisher Scientific (1 mentions)
Mir 155, by Thermo Fisher Scientific (1 mentions)
Quantstudio 7 flex real time pcr system, by Thermo Fisher Scientific (1 mentions)
Mirneasy mini kit, by Qiagen (1 mentions)
Siport neofx, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
4 protocols using mir 19b
1
Quantifying miRNA Expression in Immune Cells
2
MiRNA Transfection in Endothelial Cells
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EA.hy926 and HUVECs were seeded at 50,000/well and transfected with 100 nmol/L miRNA mimics (miR-27a/b-3p, miR-24, miR-19b or scrambled control -SCR-) from Life Technologies (Madrid, Spain) and anti-miRNA inhibitors (anti-miR-27a/b-3p or anti-SCR control) from Exiqon (Vedbaek, Denmark) using siPORT™ NeoFX™ (Life Technologies, Madrid, Spain) according to manufacturer’s instructions. After 48 h, cells and supernatants were collected for subsequent mRNA and protein analyses. Expression levels of miRNAs after transfection are shown in Supplemental Figure S6 .
3
Quantifying Circulating miRNAs by RT-PCR
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The expression of “miRNAs in the serum was identified by using quantitative real-time RT-PCR analyses and primers of circulating miRNAs, miR-328, miR-34, and miR-19b (Applied Biosystems, Foster City, CA, USA)” [79 (link)]. In this test, “GAPDH gene was applied as an internal housekeeping gene to normalize the average copy number of the resultant PCR components as previously stated in the literature” [80 (link)–82 (link)].
In PCR process, “templets of respective cDNA were subjected to four thermal phases: primary denaturation phase (I) (at 94°C for 2 minutes); denaturation phase (II) (at 94°C for 30 seconds); annealing phase (III) (at 59°C for 30 seconds); and amplification phase (IV) (at 72°C for 30 seconds)” [80 (link)–82 (link)]. “The PCR phases (II to IV) proceeded for 45 cycles, and all reactions were measured in a triplicate manner” [80 (link)–82 (link)].
In PCR process, “templets of respective cDNA were subjected to four thermal phases: primary denaturation phase (I) (at 94°C for 2 minutes); denaturation phase (II) (at 94°C for 30 seconds); annealing phase (III) (at 59°C for 30 seconds); and amplification phase (IV) (at 72°C for 30 seconds)” [80 (link)–82 (link)]. “The PCR phases (II to IV) proceeded for 45 cycles, and all reactions were measured in a triplicate manner” [80 (link)–82 (link)].
4
Establishing Gastric Cancer Cell Lines
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The human gastric adenocarcinoma cell line SGC7901 was obtained from the Academy of Military Medical Science, and MKN28 was obtained from the Shanghai Cell Bank (Shanghai, China). These two cell lines were preserved in our institute. SGC7901Luc cells stably expressing firefly luciferase were generated and preserved in our lab. All of the cells were grown in RPMI1640 (Invitrogen) supplemented with 10% heat-inactivated foetal calf serum (FCS) at 37 °C with 5% CO2 in a humidified incubator (Forma Scientific, Marietta, OH, USA). The precursors (pre-miRs) and inhibitors (miR-inhs) of the miRNAs miR-19a and miR-19b were obtained from Applied Biosystems (Invitrogen).
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