Sybr rapid quantitative pcr kit

Total RNA of leaves, roots, xylem and phloem from Poplar 84K and LBD15-oe plants were isolated by RNA Easy Fast Plant Tissue Kit (TIANGEN, Beijing, China). The cDNA was generated by reverse transcription using FastKing RT Kit (TIANGEN, Beijing, China). The qRT-PCR analysis of m⁶A pathway genes were performed with SYBR^® rapid quantitative PCR Kit (KAPA KK4601, Pleasanton, CA, USA) using the methods described previously [35 ]. Pagactin was used as a reference [24 (link)]. The primers of all the genes were listed in Table S5, and the results were analyzed using the 2^−ΔΔCt method [24 (link)].

Total RNA was isolated from the tissues of T. chinensis using the EASYspin plant total RNA isolation kit (Aidlab RN38, Beijing, China). RNA integrity and quantity were determined by 1.2% agarose gel electrophoresis and NanoDrop 1000C spectro-photometer (Thermo Scientific, Waltham, MA, USA). Reverse transcription was performed using the FastKing RT Kit (with gDNase) (TIANGEN, Beijing, China). The qRT-PCR are performed with SYBR^® rapid quantitative PCR Kit (KAPA KK4601, Pleasanton, CA, USA). Primers used for qRT-PCR are listed in Table S1. Tcactin was used as a reference gene as described previously [16 ]. Gene expression levels were calculated according to the 2^-∆∆Ct method for the different tissues and aged xylem samples [17 (link),18 (link)].

Total RNA was isolated from the tissues preserved in liquid nitrogen using the RNA extraction kit RN38 (Aidlab Biotech, China). RNA integrity was identifed with a 1.2% agarose gel, and RNA quantity and quality were determined by NanoDrop 1000C Spectro-photometer (Thermo Scientific, USA). The cDNA was obtained by reverse transcription of total RNA using the FastKing RT Kit (With gDNase) KR116 (TIANGEN, China) and used for qRT-PCR by SYBR® rapid quantitative PCR Kit (KAPA KK4601, USA). Gene-specific primers are listed in Table S2 with products lengths between 100 bp and 300 bp. Tcactin was used as a reference gene. Relative abundance of TcMYBs were calculated according to the comparative Cq method described in previous study (Li & Lu, 2014 (link); Ma et al., 2012 (link)). The expression levels of miR159, miR828 and miR858 were analyzed using the method as described previously (Shi & Chiang, 2005 (link)). One-way ANOVA was calculated using IBM SPSS 19 software. P < 0.01 was considered statistically significant and was represented by asterisks. The heat maps of differential expression of TcMYB genes were performed using HemI 1.0 with gradient bar (Clustering Method is Average linkage and Similarity Metric is Pearson distance (default)) (Deng et al., 2014 (link)).