About 1 μg genomic DNA from the R and S plants were used for DNA library preparation, using the SOLiD™ 5500 Fragment Library Core Kit (Part no. 4464412) according to its user guide. A total of 20 μg total RNA was used for two rounds of mRNA purification using Dynabeads (610.06, Invitrogen, USA). About 100 ng mRNA was fragmented using NEB Next Magnesium RNA Fragmentation Module (E6150, NEB), purified with an RNA clean up kit (R6247, Omega, USA), end repaired with T4 Polynucleotide Kinase (T4 PNK) (M0201, NEB) and cleaned up again with a kit (R6247, Omega). The end-repaired RNAs were used to prepare the strand specific transcriptome, using the Small RNA Sample Preparation kit (E6160, NEB) according to the manufacturer protocol with some minor modification, including the SR Primer F3 being replaced with barcode primers. The resulting DNA and RNA libraries were used for emulsion PCR to produce the beads for sequencing on the SOLiD 5500 machine, using 75 nt mode and 75 nt + 35 nt mode for the sequencing of DNA and RNA libraries, respectively. Biological duplicates of RNA libraries of the R and S plants before Xoo infections were profiled for quality assessment.
Small rna sample preparation kit
Manufactured by New England Biolabs
The Small RNA Sample Preparation kit is a laboratory product designed to isolate and purify small RNA molecules, such as microRNAs, from biological samples. The kit includes reagents and protocols for the extraction, purification, and preparation of small RNA samples for further analysis.
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Lab products found in correlation
Nebnext magnesium rna fragmentation module, by New England Biolabs (1 mentions)
T4 polynucleotide kinase t4 pnk, by New England Biolabs (1 mentions)
Dynabead, by Thermo Fisher Scientific (1 mentions)
Qubit 2, by Thermo Fisher Scientific (1 mentions)
Agilent 2200 tapestation, by Agilent Technologies (1 mentions)
2 protocols using small rna sample preparation kit
1
Transcriptome and Genome Profiling of Rice
2
Small RNA Sequencing Protocol
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Small RNA high-throughput sequencing was performed by Guangzhou RiboBio Co., Ltd., and the library was constructed and sequenced as follows: 1 µg of total RNA (≥50 ng small RNA) was used as the initial amount of RNA sample. The sample was supplemented with water to make the total reaction volume 7 µl. Subsequently, the library was constructed using a small RNA sample preparation kit (New England BioLabs, Inc.), according to the manufacturer's protocol. After the library was constructed, the concentration of the sample was determined by Qubit 2.0 (Thermo Fisher Scientific, Inc.), and the sample in the library was diluted to a concentration of 1 ng/µl. Library quality tests were performed using an Agilent 2200 TapeStation (Agilent Technologies, Inc.); cDNA fragments with an insert size of ~18–40 nt were obtained by gel electrophoresis.
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