Protocol PDF
0. Protocols at a glance
| PDF file | Contents | Lat updated |
| <Basic Protocols> | ||
| floatingISHe.pdf | Floating ISH with a single probe (color detction) | 2007.8.30 |
| DISHprotocol.pdf | Fluorescent double ISH | 2010.1.18 |
| PCR_cloning.pdf | Gene fragment cloning by RT-PCR | 2007.8.30 |
| <Addition> | ||
| ISHlogsheet.pdf | Logsheet for single ISH | 2007.8.30 |
| DISHlogsheet.pdf | Logsheet for double ISH | 2007.8.30 |
| <Advanced> | ||
| TSA_biotin.pdf | in prepration | |
| TISH.pdf | in prepration | |
| Tracer_ISH.pdf | in prepration | |
| BrdU_ISH.pdf | in prepration |
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1. What each protocol explains
1) Basic protocol
・floatingISHe.pdf
A protocol for floating
ISH.
Colorimetirc detection of a single probe hybridization using
NBT/BCIP substrate. Used for formalin-fixed frozen tissue.
・DISHprotocol.pdf
A protocol for fluorescent
double ISH.
In this protocol, the fluorescent detction is enabled by (1)
sensitive Alkalin phosphatase substrate, HNPP/FR and (2) amplification
of the hybridization signal using TSA (tyramide signal amplification).
They can be easily implemented for most ISH methods.
You
can find the principles and tips to use fluorescent detction.
・PCR_cloning.pdf
A protocol to construct a
probe plasmid
from RNA. Covers Reverse transcriptase (RT) PCR reaction,
subcloning of the PCR fragment into plasmid vecotrs, how to confirm the
cloned fragments, template synthesis, in vitro transcription.
2) Addition
・ISHlogsheet.pdf
Logsheet for single ISH
・DISHlogsheet.pdf
Logsheet for double ISH
・TSA_biotin.pdf
Protocol for a homemade
TSA-biotin kit.
4) Advanced
・TISH.pdf
A protocol for fluorescent
triple ISH. In preparation.
・Tracer_ISH.pdf
Tips for tracer-ISH
method. In preparation.
・BrdU_ISH.pdf
A protocol to combine
birhhtdating by BrdU and ISH. In preparation.
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