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Follow redirect from Perkin to new Revvity
AG 501-X8 Resin (Mixed bed resin for deionizing formamide), 100g, BioRad #143-7424, $323
AG 501-X8 Resin (Mixed bed resin for deionizing formamide), 100g, BioRad #143-7424, $323
[http://www.perkinelmer.com/Catalog/Product/ID/NEG034H250UC dATP-alphaS35], (250 uCi vial = 20 ul) [http://www.perkinelmer.com/AboutUs/ContactUs/ContactUS Perkin Elmer] #NEG034H250UC $172
[https://www.revvity.com/product/datp-a-thio-35s-neg034h250uc dATP-alphaS35], (250 uCi vial = 20 ul) [https://www.perkinelmer.com/corporate/contactus Perkin Elmer] #NEG034H250UC $172
:made first Tuesday of every month
:made first Tuesday of every month
'''DTT (dithiothreitol)'''
'''DTT (dithiothreitol)'''
Aka Cleland's reagent; prevents oxidation of thiol groups and breaks down disulfide bonds.
Aka Cleland's reagent; prevents oxidation of thiol groups and breaks down disulfide bonds.
==Buffer Amounts==
* Denature salmon sperm by boiling for 10 minutes.
* Put DTT in foil-covered scintillation vial.
* Add SSC, formamide, TED, salmon sperm, & Denharts solution.
* Vortex. Keep heated at 55° C on heater block until use.
<tab class="wikitable" sep="tab" border = "1" cellspacing="3" cellpadding = "3" head = top>
10 ml - - - 8 ml - - - 5 ml - - - 4 ml - - - 3 ml - - - 2 ml - -
150 mg DTT 120 mg DTT 75 mg DTT 60 mg DTT 45 mg DTT 30 mg DTT
1.0 ml SSC 20x 800 ml SSC 20x 500 µl SSC 20x 400 µl SSC 20x 300 µl SSC 20x 200 µl SSC 20x
6.0 ml Formamide 4.8 ml Formamide 3.0 ml Formamide 2.4 ml Formamide 1.8 ml Formamide 1.2 ml Formamide
2.0 ml TED 5x 1.6 ml TED 5x 1 ml TED 5x 800 µl TED 5x 600 µl TED 5x 400 µl TED 5x
1.6 ml Salmon Sperm 1.28 ml Salmon Sperm 800 µl Salmon Sperm 640 µl Salmon Sperm 480 µl Salmon Sperm 320 µl Salmon Sperm
800 µl Denharts 50x 640 µl Denharts 50x 400 µl Denharts 50x 320 µl Denharts 50x 240 µl Denharts 50x 160 µl Denharts 50x
</tab>
Note that the ceiling or maximal level of expression may vary across regions, even for the same gene product. For example, the MR has a lower and sparser expression of 5HTT and Pet-1 mRNA than the DR under baseline conditions. Thus the dynamic range of MR cells may be lower than cells in the DR, and an increase in expression within the MR may be detected as an increase in area rather than in density. The physiological bases for the heterogenity in baseline and stimulated levels of gene expression within the raphe nuclei is a profound topic, to which this paper makes a small contribution with respect to estrogen effects.
Note that the ceiling or maximal level of expression may vary across regions, even for the same gene product. For example, the MR has a lower and sparser expression of 5HTT and Pet-1 mRNA than the DR under baseline conditions. Thus the dynamic range of MR cells may be lower than cells in the DR, and an increase in expression within the MR may be detected as an increase in area rather than in density. The physiological bases for the heterogenity in baseline and stimulated levels of gene expression within the raphe nuclei is a profound topic, to which this paper makes a small contribution with respect to estrogen effects.
[[Category:Protocols]]