How to Create a Polyploid in Plants by Billy Budd

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Creating Polyploids by Billy Budd (April 20, 2013):

Abstract:

This document describes, in general, the process for creating polyploid plants with varying degrees of chromosomes. Cannabis plants are diploid in nature and are not tetraploids. Diploids have one set of chromosomes from each parent. Tetraploids have a doubled chromosome set. A tetraploid plant will grow much slower and flower slower but willl grow into giants if given enough time. The medicinal content will be signinficantly increased due to the increased size and density of the trichomes. The treatment of the dividing cells with one of these chemicals inhibits their cytokinesis during meiosis. The result is binucleate cells that contain tetraploid nuclei.

The work of polyploids done by H. E. Warmke et al. (1942-1944) suggested that the triploids were sometimes not better than either parent. I would suggest that there is no way a triploid can be stronger than a tetraploid but should be better than the diploid parent unless the offspring had a phenotype that was not well liked by the tester due to terpene composition and THC/CBD levels. We are still in the process of proving this speculation out. Triploids should be ranked right in between a diploid and a tetraploid from a purely mathematical perspective. Final data results will be available by the end of June 2013. I personally think triploids are the route to go for optimal productivity/medicinal value.

    Produce Tetraploid:

  1. There are several methods of producing tetraploids involving different chemicals including. Caffeine, Orzalin and Colchicine.
  2. The most effective method is, in my opinion, using Colchicine on the seeds and not on the plant. This is because treating the seeds will allow for the penetration of the chemical into all cells within the organism. Treating the plant will result in a polyploid that will eventually revert to diploid status. EXTREME WARNING: Colchicine is a deadly chemical and should not be used without proper ventilation, gloves and mask.
  3. DMSO (Dimethyl Sulfoxide) can be added to the solution to enhance the penetration of the chemical being employed.
  4. Prepare 3 different Colchicine solutions: .05%, .10% and .20%. This will allow for wider spectrum of experimentation but usually the .10% solution works best.
  5. Prepare three different trays of seeds for each solution strength.
  6. Soak the seeds in solution for 24 hours. Hundreds of seeds will probably be required just to make a few tetraploids.
  7. Germinate seeds in the normal fashion.
  8. Check the taproot size to determine if the polyploid conversion was successful. The taproot will be swollen compare to diploid taproots.
  9. Treatment of the seeds will result in most of the seeds either dying due to toxicity or not converting to tetraploid due to a weak solution.
  10. Fully grown the tetraploids will display the typical tetraploids features: ducksfoot, double leaf sets, bonsai appearance, giantism.

    Produce Triploid (True Sinsemilla):

  1. Produce a tetraploid plant male or female.
  2. Produce a diploid plant male if using a female tetraploid or female if using a male tetraploid.
  3. Pollinate the tetraploid female with the diploid pollen or pollinate the diploid female with the tetraploid pollen.
  4. The resulting seeds will be triploids. These are true sinsemilla and will never pollinate due to a chromosome mismatch. The plant should be medicinally stronger than a diploid but not quite as strong as a tetraploid. The triploids grow much faster than tetraploids. They are designed to be compromise between yield and super high quality. Will produce AAA product in a faster time frame than tetraploids (capable of AAAA quality).

    Produce Hexaploid:

  1. Apply a Colchicine treatment to the triploid seeds to obtain hexaploids. Hexaploids can now be mated with diploids and tetraploids to create more polyploid of varying degrees. Creating the hexaploid enables a chromosome match to facilitate such breeding.

Preparing Colchicine Solution:


From the International Carnivorous Plant Society:
Thomas Cahill
Arizona State University
Create a concentrated colchicine solution by adding 0.51 g of colchicine into 100 mL of purified water. This is a stock solution of 5100 ppm colchicine by weight. For each treatment, the stock solution was diluted to achieve the desired concentration of colchicine. For example, the 510 ppm solution involved diluting 1 mL of the stock solution by adding 9 mL of purified water in a test tube. The control group just had 10 mL of pure water added to it. The seeds were added to the test tubes and they were allowed to soak in the solution for 96 hour. The test tubes were wrapped in aluminum foil to protect the solutions from light since colchicine breaks down in light. They were stored at room temperature for the 96 hours. The solutions were shaken each day to make sure that the seeds were in contact with the solution since many of the seeds initially floated on the solution. The Sarracenia seed were stratified in a refrigerator for 4 weeks before being dosed with colchicine. NOTE: Different types of seeds will require differing concentrations and exposure times.

Preparing Colchicine Solution From Autumn Crocus Seeds (Experimental): This is my own solution due to not being able to obtain Colchicine. This is still in the experimental stage.
  1. Colchicine solutions can be prepared from the Autumn Crocus seeds
  2. Crush each of 13 seeds and add 80 mls of distilled water and soak seeds for 3-4 days at 78 degrees F or 25 C
  3. Tap root should be larger than normal
  4. Rinse with distilled water
  5. 0.5 gram for 100 mls of distilled water to make a .5% solution. Each seed is about .4 grams.
  6. Therefore mix 80 mls of distilled water with each colchicine seed if .4 grams of colchicine per seed.
  7. This method is still being proven out.

Surflan Seed Conversion Method

Kevin Vaughn Method USDA
A. Reagents
Note: ( Label all reagents with name, conc., date made, expiration date, and storage temperature. )
  1. (1) Preparation of 10?M Oryzalin solution
    • Oryzalin -99.9% pure (Molecular Weight - 346.36) Also known as 3,5-dinitro-N4N4-dipropyl-sulfanilamide. Pure reagent available from Elanco Products Co., Division of Eli Lilly & Co. Stock Solution - Using 99.9% pure Oryzalin make a 10 ?M solution in DMSO. Dissolve 0.3463 grams Oryzalin in DMSO ( Dimethyl Sulfoxide ) and dilute to 100 mls with DMSO. Working solution -10 ?M (10-5 M solution = 3.4636 mg/L) Dilute 1 ml of stock solution to 100 mls in Distilled or D.I. Water. ( 1-2 mls DMSO may be added if desired.)
  2. . (2) Preparation of 10?M Oryzalin solution using Surflan AS
    • Surflan AS - sold by Dow-Elanco as a pre-emergence herbicide under the name Surflan AS. Purity of oryzalin in surflan AS is 40.4% , thus stock solution should be adjusted accordingly.
    • Stock Solution - Using 40.4% purity Oryzalin make a 10 ?M solution in Distilled or D.I. Water. Dissolve 0.857 grams ( approx. 1 ml) Surflan AS in Distilled or D.I. Water and dilute to 100 mls.
    • Working solution -10 ?M (10-5 M solution = 3.4636 mg/L) Dilute 1 ml of stock solution to 100 mls in Distilled or D.I. Water. ( 1-2 mls DMSO may be added if desired.)
    B. Procedure
  1. Germinate seed in a container with wet filter paper or paper towels. An alternative method under less sterile conditions would be to germinate seeds on Pro-Mix or equivalent.
  2. Germinate seeds under lights - 24 hours on.
  3. Treat seeds 5-8 days from start of germination - seeds must show germination.
  4. Treat germinated seeds for 24 hours in 10 ?M working solution of Oryzalin in the dark. Use glass containers or glass petri dishes, plastic may absorb the oryzalin. Also do not use cheesecloth, as it may also absorb the oryzalin.
  5. After 24 hours the seedling roots and the base of the growing shoot should be swollen, as an indication that treatment was effective.
  6. Rinse seeds wrapped in cheesecloth or pantyhose for 4 hours under slowly running water.
  7. Transfer to promix or similar growing medium.. Plant at surface of mix, but not too deep as seed is prone to rot. Cover with plastic bag to prevent moisture loss.
  8. Cool A cool 72 F dimly-lighted area ( 1/10 th full sunlight area ) works well until good growth occurs.

How To Identify Polyploids:

You can identify a polyploid be mere physical appearance and no need to examine the chromosones. Most people have never grown or smoked these real legendary plants. The gangters rap about OG Kush because they had the money to buy this extremely rare connoisseur product. Most product out there is just "wangsta OG".
    Polyploid Traits:
  • Ducksfoot
  • Four sets of leaves at the nodes
  • Stretched (doubled) flowers
  • Dark green leaves
  • Double bud sites
  • Heavy feeder
  • High water consumption
  • Thick meristem
  • Thick leaves
  • Large stomato
  • Pistil discoloration (pink pistils under black light)
  • Extreme potency (40-45%) for tetraploids (25-35%) for triploids.
  • Vigour and extra growth on first generation (This is from research but I do not agree). I believe these plants have giant cell structure and are much slower to grow.
  • In order to correctly determine the ploidy level (number of chromosomes) then one needs to cut some roots of an actively growing plant, crush the roots, stain the roots, prepare a clean slide, put the stain crushed roots on the slide, examine under a light or electron microscope and then count the number of gametes surrounding the nuclei. This is the only correct method for determining whether a plant is a diploid, triploid, tetraplodid, etc.

EXTREME WARNING:

Colchicine is a extremely toxic alkaloid and is highly poisonous. Long term contact can result in white blood cell anomolies. You cannot smoke a plant that has been treated, post germination, with Colchicine. You can only consume the grown out product of the seeds. You can smoke product that was treated prior to germination. Selective breeding is not genetically modified cannabis. This is genetically modified pot as it was altered at the chromosonal level. Never treat a plant with Colchicine as it will become toxic and will eventually revert to diploid with further treatment. You are changing the molecular structure dynamically and none of the product can be consumed. The correct way is to treat the seeds.

Become a Master Grower

*** Read Secrets of the Pink Kush for a complete understanding of polyploids. You will learn all aspects of growing cannabis including how to grow and breed killer cannabis strains as well as other previously unknown facts about nature and giants (tetraploid humans) from this amazing research discovery.

Polyploid Strain Summary

Tetraploid marijuana plants have a real dank smell. It is really obvious when you know what you are looking for. I do not recommend growing these plants out as they take far too long to vegetate and require more attention, food and water than normal diploid plants. They could be used in a Sea of Green method but you are really relying on the genetics and not on proper plant development of hormones for optimal THC production. The resulting product is not nearly as good as a full grown plant but it is not bad for the Sea of Green method which normally produces mediocre marijuana at best. If you want to try some triploid Diablos Pink Kush seeds then check out the Seeds or get some triploid clones from me. Diablos Pink Kush is available now. Have current photos of up to week 6 vegging. Will post later.