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Pbr322 dna mspi digest ladder

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The PBR322 DNA-MspI Digest ladder is a pre-prepared DNA fragment size standard used for size determination of DNA samples during gel electrophoresis. It consists of DNA fragments of known sizes created by digesting the pBR322 plasmid with the MspI restriction enzyme.

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Lab products found in correlation

Hiseq 2500 machine, by Illumina (3 mentions) Zr small rna ladder, by Zymo Research (2 mentions) Hiseq 2500, by Illumina (1 mentions)

Close spelling variants of this product

PBR322 DNA (2 citations) DNA ladder (35 citations) PBR322 (16 citations)

4 protocols using pbr322 dna mspi digest ladder

1

Small RNA Sequencing from Floral Buds

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4 un-opened flower buds (stage 12 and younger) from individual
mutant plants as well as 3 individual wild-type (WT) controls were
collected, frozen in liquid nitrogen and kept at −80°C until
use. The total RNA extraction and small RNA enrichment were performed as
previously described68 (link)with the following minor modifications: (1) for the small RNA enrichment
step an equal volume of 20% polyethylene glycol 8000/2M NaCl was added to
each total RNA sample and (2) the ZR-small RNA ladder (Zymo Research, Cat#
R1090) was used to determine the gel region corresponding to the
17–29 nucleotide (nt) size range. The resulting small RNAs were then
used for library preparation with the NEBNext Multiplex Small RNA Library
Prep Set for Illumina (New England Biolabs, Cat# E7300) following the
user’s manual. The final library products were further purified using
an 8% polyacrylamide gel to excise 130–160nt products relative to the
pBR322 DNA-MspI Digest ladder (New England Biolabs, Cat# E7323AA). The
libraries were pooled and sequenced (single end 50bp, SE50) on a HiSeq 2500
machine (Illumina).
Zhou M., Palanca A.M, & Law J.A. (2018). Locus-specific control of the de novo DNA methylation pathway in Arabidopsis by the CLASSY family. Nature genetics, 50(6), 865-873.
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2

Small RNA Sequencing from Plant Tissues

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Total RNA extraction and small RNA enrichment were performed as previously described in Zhou et al.37 . Briefly, 2.0 µg of total RNA from rosette and leaf tissues, or ~0.5 µg of total RNA from ovules, were used for smRNA enrichment. The resulting small RNAs were used for library preparation with the NEBnext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs, Cat# E7300) following the user’s manual. The final library products were further purified using an 8% polyacrylamide gel to excise 130-160nt products relative to the pBR322 DNA-MspI Digest ladder (New England Biolabs, Cat# E7323AA). The purified libraries were pooled and sequenced (single end 50 bp, SE50) on a HiSeq 2500 machine (Illumina).
Zhou M., Coruh C., Xu G., Martins L.M., Bourbousse C., Lambolez A, & Law J.A. (2022). The CLASSY family controls tissue-specific DNA methylation patterns in Arabidopsis. Nature Communications, 13, 244.
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3

Small RNA Sequencing from Floral Buds

Check if the same lab product or an alternative is used in the 5 most similar protocols
4 un-opened flower buds (stage 12 and younger) from individual
mutant plants as well as 3 individual wild-type (WT) controls were
collected, frozen in liquid nitrogen and kept at −80°C until
use. The total RNA extraction and small RNA enrichment were performed as
previously described68 (link)with the following minor modifications: (1) for the small RNA enrichment
step an equal volume of 20% polyethylene glycol 8000/2M NaCl was added to
each total RNA sample and (2) the ZR-small RNA ladder (Zymo Research, Cat#
R1090) was used to determine the gel region corresponding to the
17–29 nucleotide (nt) size range. The resulting small RNAs were then
used for library preparation with the NEBNext Multiplex Small RNA Library
Prep Set for Illumina (New England Biolabs, Cat# E7300) following the
user’s manual. The final library products were further purified using
an 8% polyacrylamide gel to excise 130–160nt products relative to the
pBR322 DNA-MspI Digest ladder (New England Biolabs, Cat# E7323AA). The
libraries were pooled and sequenced (single end 50bp, SE50) on a HiSeq 2500
machine (Illumina).
Zhou M., Palanca A.M, & Law J.A. (2018). Locus-specific control of the de novo DNA methylation pathway in Arabidopsis by the CLASSY family. Nature genetics, 50(6), 865-873.
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4

Small RNA Sequencing Protocol

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Inflorescences from three biological replicates of WT, zmp-2, nrpd1-3, pZMP::ZMP-YFP/zmp-2, zmp-2 nrpd1-3, and zmp-2 shh1-1 were collected and frozen in liquid nitrogen and kept at −80°C until use. Total RNA was extracted as described above and resolved in 15% urea–PAGE gels, from which gel pieces corresponding to the 15- to 40-nt sRNA fraction were excised. The sRNAs were recovered by soaking the gel slices in 0.4 N NaCl, followed by ethanol precipitation. The resulting sRNAs were then used for library preparation with the NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs, E7300) following the user’s manual. The final library products were resolved on a 12% UREA-PAGE gel, from which the 150-bp band as determined by the pBR322 DNA-MspI Digest ladder (New England Biolabs, E7323AA, provided in New England Biolabs 7300) was excised. The libraries were pooled and sequenced (single-end 50-bp, SE50) on a HiSeq 2500 instrument (Illumina).
Wang Y., Le B.H., Wang J., You C., Zhao Y., Galli M., Xu Y., Gallavotti A., Eulgem T., Mo B, & Chen X. (2022). ZMP recruits and excludes Pol IV–mediated DNA methylation in a site-specific manner. Science Advances, 8(47), eadc9454.
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