2
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9
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Easy to use, add 2ul per 10ul of DNA solution.
1
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27
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Basic protocol for Eosin & Haematoxylin histological preparation of microdissection slide.
1
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46
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Paper strip method for DNA extraction from agarose gels
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45
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Ligation of a DNA fragment to a vector
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16
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Isolate a DNA fragment from Low Melting agarose
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30
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The removal of phosphate groups from DNA using Alkaline Phosphatase
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64
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Calculate DNA concentration for efficient DNA ligations.
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24
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One way to make transgenic animals in C. elegans we use a microinjection technique. Briefly, a DNA construct (plasmid, cosmid or YAC) or PCR product with your gene(s) of interest is mixed with a co-injection marker and injected into the distal gonad (syncytium). The injected DNA is taken up into the mature oocyte's nucleus. The DNA exists as an extrachromosomal array (i.e. not integrated in the chromosome) which segregates randomly and can be lost, that is why we need a marker to follow which animals have the array. F1 progeny that show the co-injection marker phenotype are picked and transgenic lines are established by keeping those animals that segregate the array in their F2 generation. Usually 1 in 10 F1 progeny with the array will give you a transgenic line.
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DNA Extraction via Chelex
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The purpose of this procedure is used to chew up excess primers and remove excess
dNTPs from your PCR product. This proceedure is necessary to ensure clean and
readable DNA sequences. Work in HIGH DNA part of
lab.
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21
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In this protocol a biotin or digoxigenin labelled DNA probe is detected using HRP-conjugated antibodies. The signal is visualised with diaminobenzidine (DAB). Normal, healthy nuclei show two spots. Aneusomic nuclei show 1, 3, 4 or more spots.
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18
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Transformation of plasmid DNA to competent E. Coli cells
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PCR mutagenesis is a method for generating site-directed mutagenesis. This method can generate mutations (base substitutions, insertions, and deletions) from double-stranded plasmid without the need for subcloning into M13-based bacteriophage vectors and for ssDNA rescue.
This method uses a proof-reading polymerase to read all the way around a plasmid and thus incorporate the primer as the new (mutant) sequence. Only a few (say 12) PCR cycles are performed on a relatively large amount of plasmid template to minimise the chance of expanding PCR sequence errors.
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23
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These are excellent for extracting DNA if you can afford them. They cost 1–2 US$ each. Manufacterers include Qiagen, Sigma, Novagen
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58
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Dialysis tubing (semi-permeable membrane, Visking tubing)
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95
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Preparation of Sonicated Salmon Sperm DNA
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11
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C. elegans total genomic DNA preparation
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7
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This is an example of how to measure the lengths of DNA contours on images acquired using an atomic force microscope (AFM).
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5
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A Salting Out Procedure for DNA extraction from cheek cells obtained by rinsing the mouth with 25mls of any commercial mouth wash solution available for about 30sec the first thing in the morning. It is important not to brush your teeth before collecting the specimen.