Zr plant rna miniprep kit

Total RNA was isolated from roots using a ZR Plant RNA MiniPrep kit (Zymo research). The RNA concentration was estimated spectrally (Nano Drop ND-1000; Nano Drop Technologies). cDNA was synthesized using a qScript cDNA synthesis kit (Quanta). The reaction mixture contained 700 ng total RNA and random primers. Primer design and RT-qPCR assays for determining relative steady-state transcript levels were as previously described [17 (link)]. Primers are described in Table S1.

Total RNA was isolated from C. kanran buds and flowers using the ZR Plant RNA MiniPrep Kit (Zymo Research, Irvine, CA, USA), and 1 μg RNA was used to synthesise oligo(dT)-primed first-strand cDNA by the QuantiTect Reverse Transcription Kit (QIAGEN, Hilden, Germany) following the protocol of the manufacturer. The qRT-PCRs were performed in a 7300 Real-Time PCR System (Applied Biosystems, Foster Arthritis 5 City, CA, USA) using an SYBR premix Ex TaqTM Kit (TaKaRa, Kyoto, Japan). The reference gene we chose was actin (c114989.graph_c0). We calculated the relative expression level by the 2-^ΔΔCT method (Livak and Schmittgen, 2001 (link)).

Total RNA was extracted using the ZR Plant RNA MiniPrep kit (Zymo Research, Irvine, CA, USA) following the manufacturer's protocol. The quality of the extracted RNA was evaluated by electrophoresing on 1% agarose gel. The concentration was determined with a Qubit 2.0 fluorometer (Life Technologies, Carlsbad, CA, USA) as described in the manufacturer's protocol.

Three pools of leaves, each of which contained five control or stressed plants of the two studied genotypes, were used for total RNA isolation. The sampling was done at 0, 2, 8, and 24 h after salt stress application [91 (link)]. RNA extraction was performed using the ZR Plant RNA MiniPrep™ Kit (Zymo Research, Irvine, CA, USA). DNA was eliminated from RNA samples by TURBO DNA-free™ Kit (Promega, Madison, WI, USA). The quality and quantity of the isolated RNAs were checked by agarose gel electrophoresis (1%) and spectrophotometrically using BioPhotometer (Eppendorf BioPhotometer plus, Hamburg, Germany).
The three replicates of each treatment for both genotypes were sequenced at the Beijing Genomics Institute (BGI, Shenzhen, China) using the Illumina NextSeq 500 platform as described in our previous publication [91 (link)].
The clean sequencing reads were submitted to SRA database at NCBI (accession number: PRJNA821484).

Total RNA was isolated from shoots and roots representing each time point using the ZR Plant RNA MiniPrep™ Kit (Zymo Research, Irvine, CA, USA). The quality and quantity of isolated RNAs were checked by agarose gel electrophoresis and spectrophotometrically using a BioPhotometer (Eppendorf BioPhotometer plus, Hamburg, Germany). Residual DNA was eliminated using a TURBO DNA-free™ Kit (Promega, Madison, WI, USA).

Young seedlings were collected 14 days after germination and leaf blades, sheath, stems, and panicles were collected 60 days after germination. Total RNA were isolated from frozen tissues using a ZR Plant RNA MiniPrep Kit (ZYMO Research, Beijing, China) and reverse transcribed using a QuantiTect reverse transcription kit (Qiagen, Shanghai, China) according to the manufacturer’s protocol. RT–PCR was performed with an SYBR premix Ex Taq Kit (TaKaRa, Kusatsu, Japan) according to the operation manual and amplified in an ABI 7500 using primers listed in Supplementary Table S5. Data from three biological replicates were analyzed following the ΔΔCT method.