Navigation Links
DNA Isolation From 300 to 450 mg Dried or 600 to 1,200 mg Fresh Leaf Tissue


AquaPure Genomic DNA Isolation Kit
Catalog #732-6340
Expected yield range: 90900 g DNA


Method
Cell Lysis
1. Add 300-450 mg dried tissue (finely ground) or 6001,200 mg frozen tissue (which may be finely ground with a mortar and pestle in liquid nitrogen), or 6001,200 mg fresh tissue to a 50 ml tube. Work quickly and keep tissue cold to minimize DNase activity. Note: it may be necessary to vary the amount of starting material depending upon the species, age, tissue preparation, and genome size.

2. Add 18 ml cell lysis solution to the leaf tissue. For dried tissue, cap the tubes and vortex 13 sec to wet the tissue. For unground tissue, homogenize using 3050 strokes with a tube pestle.

3. Incubate cell lysate at 65C for 60 min. After 30 and 60 min, invert capped tube 10 times.

RNase Treatment (Optional)
1. Add 90 l RNase A solution to the cell lysate.

2. Mix the sample by inverting the capped tube 25 times and incubate at 37C for 15 min.

Protein Precipitation
1. Cool sample to room temperature.

2. Add 6 ml protein precipitation solution to the lysate.

3. Cap the samples and vortex vigorously at high speed for 20 sec to mix the protein precipitation solution uniformly with the cell lysate. Alternatively, invert a rack containing the samples 150 times (approximately 2 min) to mix the protein p recipitation solution uniformly with the cell lysate. For species with high polysaccharide content, it may be necessary to incubate sample on ice for 515 min.

4. Centrifuge at 3,000 x g for 10 min. The precipitated proteins should form a tight, green pe llet. The supernatant may range in appearance from a transparent brown to green, depending on the sample. If the pellet is not tight, repeat step 3, incubate on ice for 5 min, then repeat step 4.

DNA Precipitation
1. Leaving behind the precipitated protein pellet, pour the supernatant containing the DNA into a 50 ml centrifuge tube that contains 18 ml 100% isopropanol (2-propanol).

2. Mix the sample by inverting the capped tube gently 50 times.

3. Centrifuge at 3,000 x g for 5 min. The DNA will be visible as a pellet that ranges in color from off-white to dark green, depending on particulates that cosediment with the DNA.

4. Pour off supernatant and drain tube briefly on clean absorbent paper. Add 18 ml 70% ethanol and invert the capped tube several times to wash the DNA pellet.

5. Centrifuge at 3,000 x g for 5 min. Carefully pour off the ethanol. Pellet may be loose, so pour slowly and watch the pellet to ensure that it stays in the tube.

6. Invert and drain the tube on clean absorbent paper and allow to air-dry 15 min.

DNA Hydration
1. Add 1 ml DNA hydration solution.

2. Rehydrate DNA by incubating sample 1 hr at 65C, or a l t e rn a t i v e l y, overnight at room temperature. Tap tube periodically to aid in dispersing the DNA. Transfer DNA sample to a clean 1.5 ml tube for storage.

3. If particulates are present in the rehydrated DNA sample, centrifuge at 13,00016,000 x g for 510 min and transfer supernatant into a clean 1.5 ml tube. Store DNA at 28C.


back to top
'"/>

Source:


Page: All 1 2

Related biology technology :

1. Fast and Easy Isolation of PCR-Ready Genomic DNA from Whole Blood
2. High-Throughput Isolation of Total RNA
3. Simple Isolation of RNA from Tissue and Cultured Cells
4. Fast Isolation of Total RNA from Small Numbers of Cells
5. Comparison of Growth Techniques and Media for the Purpose of Plasmid Isolation from E. coli Using the Eppendorf Perfectprep Plasmid Mini Kit
6. Isolation of Microorganisms
7. Isolation of Genomic DNA from Saliva Using the Perfect gDNA Blood Mini Kit
8. Mouse Tail Genomic DNA Isolation Protocol(1)
9. Genomic DNA Isolation Protocol(1,2,3)
10. Basic Plasmid DNA Isolation Protocol
11. Protocol for RNA Isolation Using TRIzol Reagent with Phase Lock Gel-Heavy
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:10/10/2017)... 10, 2017 SomaGenics announced the receipt of ... develop RealSeq®-SC (Single Cell), expected to be the first ... (including microRNAs) from single cells using NGS methods. The ... to accelerate development of approaches to analyze the heterogeneity ... "New techniques for measuring levels of mRNAs in individual ...
(Date:10/9/2017)... 2017  BioTech Holdings announced today identification and ... ProCell stem cell therapy prevents limb loss in ... demonstrated that treatment with ProCell resulted in more ... compared to standard bone marrow stem cell administration.  ... reduction of therapeutic effect.  ...
(Date:10/9/2017)... ... October 09, 2017 , ... ... line of medical marijuana products targeting the needs of consumers who are incorporating ... Kindred takes place in Phoenix, Arizona. , As operators of two successful Valley ...
(Date:10/7/2017)... ... October 06, 2017 , ... ... launched its ProxiMeta™ Hi-C metagenome deconvolution product, featuring the first commercially available ... bioinformatics software to perform Hi-C metagenome deconvolution using their own facilities, supplementing ...
Breaking Biology Technology:
(Date:4/11/2017)... No two people are believed to ... York University Tandon School of Engineering and Michigan ... partial similarities between prints are common enough that ... and other electronic devices can be more vulnerable ... in the fact that fingerprint-based authentication systems feature ...
(Date:4/5/2017)... NEW YORK , April 5, 2017 ... security, is announcing that the server component of the ... is known for providing the end-to-end security architecture that ... customers. HYPR has already secured over 15 ... system makers including manufacturers of connected home product suites ...
(Date:3/30/2017)... ANGELES , March 30, 2017  On April ... Hack the Genome hackathon at Microsoft,s ... exciting two-day competition will focus on developing health and ... Hack the Genome is the ... been tremendous. The world,s largest companies in the genomics, ...
Breaking Biology News(10 mins):