Section2_replica4_techrep2_p
Three zones were sampled, from the base to 3.5 cm, from 3.5 to 7.0 cm and from 7.0 to 10.5 cm, of the developing fourth leaf of maize B104 plants at two days after leaf appearance (DALA). Only leaf samples were taken from well-watered plants.    To sample the three zones of a developing fourth leaf two days after leaf emergence, the older leaves that surround the growing leaf should be removed. Special care should be taken not to damage the basal part of the growing leaf because this is where the leaf basal meristem is located. Subsequently the fourth leaf was cut into three parts corresponding to the three sampled zones.  B104 seeds were sown in soil and grown in a controlled-environment growth chamber (relative humidity (55%), temperature (24C day/18C night) and 16 h/8 h (day/night) cycle). Only well-watered plants.  Purified poly(A)-containing mRNA molecules were fragmented, randomly primed strand-specific cDNA was generated and adapters were ligated.  Total RNA was extracted using the guanidinium thiocyanate-phenol-chloroform extraction method using TRI-reagent (Thermo Fisher Scientific) followed by DNA digestion using the RQ1 RNase-free DNase kit (Promega). Library preparation was done using the NEBNext Kit (Illumina).