Germinate weed seeds peroxide

Development and Standardization of Rapid and Efficient Seed Germination Protocol for Cannabis sativa

Cannabis seed germination is an important process for growers and researchers alike. Many biotechnological applications require a reliable sterile method for seed germination. This protocol outlines a seed germination procedure for Cannabis sativa using a hydrogen peroxide (H2O2) solution as liquid germination media. In this protocol, all three steps including seed sterilization, germination, and seedlings development were carried out in an H2O2 solution of different concentrations; 1% H2O2 solution showed the fastest and the most efficient germination. This protocol also exhibited high germination efficiency for very old cannabis seeds with lower viability. Overall, this protocol demonstrates superior germination compared to water control and reduces the risk of contamination, making it suitable for tissue culture and other sensitive applications.

Keywords: Cannabis sativa, Rapid germination, Hydrogen peroxide, Seed sterilization, Seedling development

Background

Cannabis sativa, otherwise known as marijuana or hemp, is an annual primarily dioecious flowering plant in which male/female sex is determined by heteromorphic chromosomes (X and Y) ( Gaudet et al., 2020 ). Cannabis is grown for a variety of agricultural uses; nearly all parts of cannabis plant are used, seeds for food, stem for fiber, and flowers/leaves for medicine. Flowers produce a mix of cannabinoids and aromatic compounds valued for their therapeutic and recreational effects ( Chandra et al., 2017 ). Cannabis plants are propagated either clonally through cuttings or via seed germination. Seed germination is very important for researchers, breeders, and growers alike, especially since seeds from elite cultivars can be very expensive and valuable. Additionally, older seeds may have a reduced germination rate while bacterial and fungal contamination can compromise germination, especially when seeds are germinated for tissue culture propagation. To address these issues, we have developed a rapid, sterile, and efficient seed germination protocol using a 1% hydrogen peroxide (H2O2) solution. In this protocol, all three steps including seed sterilization, germination, and seedlings development were carried out in a 1% H2O2 solution. This presents a significant advantage over other sterilants, such as mercuric chloride or bleach, which require additional washing of seeds and a separate germination step on MS solid medium. Our protocol resulted in faster germination and increased seed germination percentage as compared to water control, with no bacterial or fungal contamination, making it suitable for tissue culture and other sensitive applications. In comparison to previous germination methods which take between 4-7 days for radicle appearance and 5-15 days for seedling development ( Wielgus et al., 2008 and references therein), our germination method resulted in radicle appearance in 1 day and allowed us to obtain cannabis seedlings in a very short period (3-7 days) with minimal efforts. This protocol is also very efficient for germination of very old cannabis seeds with lower viability.

Materials and Reagents

All seeds were harvested in our laboratory. Blueberry seeds were not older than 6 months, when employed in the experiments. Finola and X59 seeds were more than 5 years old.

Equipment

Growth chamber (Sanyo MLR-350, catalog number: 859-600-06): 24 °C, 18 h light/6 h dark cycle, light intensity 200 μmol·m -2 ·sec -1

Procedure

Seed germination assay

Soak seeds overnight in various concentrations of hydrogen peroxide solution (liquid germination media or germination solutions) as well as in sterile water control (H2O, 1% H2O2, 3% H2O2, 5% H2O2, or 10% H2O2) in 15 or 50 ml screw-cap (Falcon tube). Falcon tubes with submerged seeds in various germination solutions were kept in the dark at room temperature.

Next day, record the percentage of germinated seeds in germination solution (appearance of radicle is considered as germination event) and add fresh respective germination solution after removal of old solution simply by pouring out.

Keep seeds soaked in the same solution for 3 more days in the dark at room temperature and record the percentage of germinated seeds every day.

Thereafter, germinated seeds/seedlings were transferred with or without seed coats from H2O2 solution to MS medium plates to observe the growth of H2O2 solution-germinated seeds/seedlings on MS medium. To transfer, first germinated seeds/seedlings were poured together with H2O2 solution from the Falcon tube to the empty petri plate. Then seedlings were transferred to sterile paper by using forceps to remove excess H2O2 solution. Finally, the germinated seeds/seedlings were transferred to MS media plate by using forceps. The whole transfer process has been carried out in the laminar flow hood.

Parafilm sealed MS medium plates with germinated seeds/seedlings are then transferred to the growth chamber (24 °C, 18 h light/6 h dark cycle and light intensity 200 μmol·m -2 ·sec -1 ) for 3 days to observe the growth and survival of H2O2 solution germinated seeds/seedlings on MS medium.

The H2O2 solution-germinated seeds/seedlings growth was also observed in soil. Pro-Mix HP Mycorrhizae Growing Medium used for soil experiment. The cannabis seeds were soaked in the H2O2 solution (germination solutions) for four days and thereafter, germinated seeds/seedlings were transferred from H2O2 solution to soil pot (Pro-Mix HP Mycorrhizae Growing Medium) to observe the growth and survival of H2O2 solution germinated seeds/seedlings on soil. The soil pots were transferred to the growth chamber (24 °C, 18 h light/6 h dark cycle and light intensity 200 μmol·m -2 ·sec -1 ). The photographs were taken on day 12.

Data analysis

Mean seed germination percentage under various concentrations of H2O2 solution as well as water control were calculated in an excel sheet. Data were shown as mean ± SE.

Results

In this study, we have described a rapid and efficient seed germination protocol for Cannabis sativa. The brief description of this protocol has been reported in Sorokin et al. (2020) . In the current study, we have standardized the optimum concentration of hydrogen peroxide (H2O2) solution media for efficient sterilization and rapid germination. We have tested various concentrations of H2O2 solution as well as sterile water control (H2O, 1% H2O2, 3% H2O2, 5% H2O2, or 10% H2O2) for sterilization and germination efficiency. All three steps of germination (seed sterilization, germination, and seedlings development) were carried out in various concentrations of H2O2 solution and seeds were kept in liquid media for four days. Hydrogen peroxide presents several significant advantages over mercuric chloride or bleach sterilants, which require additional seed washing, and separate germination/seedling development step in Murashige and Skoog (MS) agar medium ( Sorokin et al., 2020 ). The 1% H2O2 solution showed rapid and higher germination than higher H2O2 concentrations solution and water control at day 1 ( Figure 1 ). On day 1, 1% H2O2 solution exhibited 82.5% germination as compared to 22.5% germination for 3% H2O2 group, 17.5% germination for 5% H2O2 group and 47.5% germination in water control group ( Figure 1B ). Interestingly, 10% H2O2 did not show any germination on day 1 due to its toxic effect ( Figure 1 ). In 1% H2O2 solution, radicle appearance (germination) occurred within 24 h and seedling development (two fully developed cotyledons and two immature true leaves stage) occurred in 72-96 h ( Figure 1A ). In comparison to previous germination methods which take between 4-7 days for radicle appearance and 5-15 days for seedling development ( Wielgus et al., 2008 and references therein), our germination method resulted in radicle appearance in 1 day and allowed us to obtain cannabis seedlings in a very short period (3-7 days) with minimal efforts ( Figures 1 -2). Considering the possible toxic effect of H2O2 (since germinated seeds/seedlings stayed continuously in H2O2 solution for 4 days), we have checked further survival of germinated seeds/seedlings on MS media and soil ( Figures 2 -3). On MS media, 1% H2O2 solution seedlings survived better than other treatments ( Figure 2 ). The water germinated seeds exhibited contamination and did not survive on MS media ( Figure 2 ). Similarly, due to the toxic effect of higher concentration of H2O2, the 10% H2O2 germinated seeds did not survive on MS media ( Figure 2 ). The 1% H2O2 solution seedlings also survived well on soil ( Figure 3 ). Apart from this, we have also tested our method for more than 5-years old cannabis seeds with lower viability, which demonstrated that 1% H2O2 solution medium exhibited a very high germination percentage (~50%) as compared to water control (~10%) ( Figure 4 ). In conclusion, we have developed a rapid and efficient method for C. sativa seed germination under sterile conditions for tissue culture and other sensitive applications.

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Germination of 6-month-old seeds of Blueberry variety in various concentrations of hydrogen peroxide solution and water control.

A. Representative photographs of germinated seeds/seedlings in the H2O2 solution of various concentrations or water control on day 1 to day 4. B. Comparison of germination percentage between the various concentrations of H2O2 solution or water control. Data are shown as mean ± SE (n = 4). In each replicate, 30 seeds were used.

Representative photographs of growth and survival of H2O2 solutions germinated seeds/seedlings of Blueberry variety on MS media.

The Blueberry variety seeds were soaked in the H2O2 solution (germination solutions) for four days and thereafter, germinated seeds/seedlings were transferred from H2O2 solution to MS medium plates to observe the growth and survival of H2O2 solution germinated seeds/seedlings on MS medium. The photographs were taken at day 0 (just after transfer to MS medium plates), day 1 (after 24 h of the transfer to MS medium plates), and day 3 (after 72 h of the transfer to MS medium plates) on MS media.

Representative photograph of Blueberry variety young plantlet growing in soil (Pro-Mix HP Mycorrhizae Growing Medium).

The Blueberry variety seeds were soaked in the H2O2 solution (germination solutions) for four days and thereafter, germinated seeds/seedlings were transferred from H2O2 solution to soil pot (Pro-Mix HP Mycorrhizae Growing Medium) to observe the growth and survival of H2O2 solution germinated seeds/seedlings on soil. The photographs were taken on day 12.

Germination of 5-years old seeds of Finola and X59 varieties in 1% hydrogen peroxide solution and water control.

Comparison of germination percentage between 1% H2O2 solution media and water control. Data are shown as mean ± SE (n = 5). In each replicate, around 30 seeds were used.

Recipes

4.43 g Murashige & Skoog Basal Medium with Vitamins

Adjust pH to 5.7 with KOH and sterilize by autoclaving at 121 °C for 40 min. 25 ml of MS media on each Petri plate.

Acknowledgments

This protocol is derived from Sorokin et al. (2020). We thank the Natural Sciences and Engineering Research Council of Canada (NSERC) and MITACS for funding our work.

Competing interests

The authors declare that they have no competing interests.

Citation

Readers should cite both the Bio-protocol article and the original research article where this protocol was used.

References

1. Chandra S., Lata H. and ElSohly M. A.(2017). Cannabis sativa L.-botany and biotechnology. Chandra, S., Lata, H. and ElSohly, M. A.(Eds.). Springer International Publishing: Cham, Switzerland. ISBN: 9783319545639. [Google Scholar]

2. Gaudet D., Yadav N. S., Sorokin A., Bilichak A. and Kovalchuk I.(2020). Development and optimization of a germination assay and long-term storage for Cannabis sativa pollen . Plants 9 : 665. [PMC free article] [PubMed] [Google Scholar]

3. Sorokin A., Yadav N. S., Gaudet D. and Kovalchuk I.(2020). Transient expression of the β-glucuronidase gene in Cannabis sativa varieties . Plant Signal Behav 15 ( 8 ): 1780037. [PMC free article] [PubMed] [Google Scholar]

4. Wielgus K., Luwanska A., Lassocinski W. and Kaczmarek Z.(2008). Estimation of Cannabis sativa L. tissue culture conditions essential for callus induction and plant regeneration . J Nat Fibers 5 : 199-207. [Google Scholar]

Germinating Marijuana Seeds: Step by Step Guide

The life of any plant begins with germination , making this process the most important in your plant’s life. You will undoubtedly come across many different methods for germinating your marijuana seeds when conducting your online research. Some may be successful, but others will disappoint you. The following guide will provide you with an effective list of practices and critical information home marijuana growers can utilize to help ensure they’re preparing the best crop possible to produce some smooth, enjoyable hits.

Please continue reading to find out everything you need to know about cannabis seed germination. And if you’d like even more information on the wonders of growing cannabis from the comfort of your own home, please consider exploring the top-quality grow kits provided by our cannabis industry experts at a Pot for Pot today , along with their range of expertly written educational articles.

What Does it Mean to Germinate Cannabis Seeds?

Germination is the natural process by which a particular organism grows from a spore or seed. In this case, the term is referring to the sprouting of a cannabis seedling from a seed , which can be encouraged by help from the cannabis grower. Germinating your seed is the first crucial step in the cannabis growing process, and it can be surprisingly simple with the right tools. Even though the process of getting a seed to sprout and bear a healthy plant is a challenge for some growers, it doesn’t have to be when you start with a Pot for Pot. With our help, your plant will sprout and start growing in no time!

What to Understand About Germination and Endosperm

The first thing to understand is that seeds come naturally equipped with their own starting food- endosperm. It plays an essential role in ensuring your plant reaches its full potential. As a seed begins to sprout, this starchy blanket around the embryo provides nutrients to a growing plant. If it doesn’t seem like a lot, don’t be fooled- there’s enough starch there to feed humans.

That starch is only part of what you’ll need, however. You’ll also need to provide specific temperatures to germinate your seeds. Ideally, you’ll want to keep the temperature around 25 degrees Celsius (77 F) for the best results.

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What Happens During Germination?

The process of germination is when plants start to sprout from their seeds, and it’s relatively the same throughout the plant kingdom. A little bit of moisture makes the tiny plant inside crack open its outer shell and sprout upwards while pushing its root downwards. Once it’s cracked through the shell, you’ll need to give your little babies a bit of water and some love for the fun to begin.

Your seeds will need the right conditions to break through their shell and continue to grow, however. Otherwise, they may take longer to germinate. Our seedling starter kit can help you create those perfect conditions. It makes sure your seed has just the right amount of water (so that it stays moist, but not too wet). Once it has germinated and grown enough to sprout its third set of leaves, simply move your young plant into the Pot for Pot fabric pot for continued healthy growth.

It’s best to germinate seeds indoors where you can better control the temperature. Seeds are more likely to sprout in a warm and stable environment.

How to Sprout Marijuana Seeds

Marijuana plants start as a seed . That tiny thing resembling a pebble is a whole plant securely stored with a supply of food to support itself for a few days. In the germination process, the food converts into sugars, which the plant uses to cut the shell and develop its root. Identifying a healthy seed is, therefore, the first thing that you need to do.

The sprouting process is the foundation of your marijuana plant, so do not take this step lightly. Other than starting with a high-quality seed, you need to ensure that your conditions are optimal. It is challenging to optimize the natural mediums, which is why a medium such as Jiffy Pellets comes in handy.

Different Ways to Germinate

There are many ways to germinate seeds, but we recommend using peat pellets. Not only is it the safest way to start your seeds, but it’s also the easiest. Jiffy is hands down one of the best brands of peat pellets on the market , and if you are thinking about germinating using peat pellets, this is the way to go. This method is great for avoiding the risk of damaging new roots. We’ll get into more detail about the wonders of Jiffy Pellets in a later section of this article. But first, let’s explore the benefit of peat pellets in general, along with some other mediums growers can use to facilitate effective cannabis germination.

Peat pellets are comprised of decomposed vegetable matter which your plants will absolutely love. When you add water, it expands, creating a nutrient-dense medium that is a good replacement for soil. When the roots are visible, you can transport the whole pellet into a Top Soil mix with fortified Pot for Pot Superb Soil so it can continue developing without accidentally causing root damage.

Basic Mediums for Marijuana Germination

There are many different germination methods, and results will vary. Here are some basic ways to sprout your seeds :

Sprouting marijuana seeds in regular soil is a common practice. Soil is a natural medium and can protect the seeds as they develop their fragile roots. For this option, be sure to start with the right type of soil for marijuana. Use a seed starter or fertilized potting soil with a pH of around 6. Soil has both minerals and spores that support growing marijuana plants.

Water

Water is another method for sprouting a marijuana seed. This method produces a seedling in 24-48 hours. The advantage of using water is that you can ensure adequate moisture for germination. The plant will also have an easier time breaking its shell compared to when it is in a soil medium. For this method, water daily while checking the temperature, ensuring that it is around 65° F.

Rockwool

This mineral wool comes from volcanic rock and other materials such as limestone and basalt. To make this material, the ingredients are heated into molten lava and rapidly spun into threads. These threads then undergo compaction, curing, and finally, cutting.

While Rockwool is a suitable environment for germination, you’ll need to adjust it a bit for growing marijuana. First, you’ll need to add fertilizers- around 600 parts per million. You’ll also need to reduce the pH, which is too high to support sprouting. It’s also a bit dangerous to use, so please wear gloves and protect your mouth and eyes while handling it.

Would you like to save money and reduce the number of trips you need to take to your local dispensary to stock up on marijuana? Consider trying out the potential of your green thumb by investing in one of our top-quality cannabis growing kits today.

The Best Marijuana Germination Medium: What to Know About Jiffy Pellets

For the best results, you need a precise understanding of how to get started growing your marijuana plant. As stated a few sections above in this article, choosing Jiffy Pellets as your initial medium is probably the best way to go. Every Pot for Pot kit includes a Jiffy Pellet, so you don’t need to worry about purchasing it separately, and if you run out, you can order some refills or one of our expansion kits .

What is a Jiffy Pellet and How Does it Work?

A Jiffy Pellet is a natural, biodegradable growing medium conveniently available as little compact discs that help sprout seeds into seedlings. Jiffy Pellet composition includes peat moss, a mesh, and other ingredients such as lime, fertilizer, and ammonium to help seedlings grow.

Jiffy Pellets are both a container and a potting mix for sprouting seeds. Peat moss is a medium that’s rich in organic materials, and the mesh securely holds the peat moss in place. Using Jiffy peat pellets either indoors or outdoors is an effective method to grow healthy and robust marijuana plants.

To use a Jiffy pellet, water it first. Then, place your marijuana seed a bit below the surface level. When the plant is strong enough, transplant the entire pellet to a larger pot.

What are Jiffy Pellets Composed of?

To help you better understand the advantages that Jiffy Pellets can provide to your seed germination efforts, let’s take a moment to explore the various beneficial components they contain and how they work to benefit your plants.

Technically, a Jiffy peat pellet is a small disc made of compressed material. However, Jiffy pellets enlarge up to seven times their original size when you add water , creating a suitable environment for the sprouting process. The fine netting keeps the Jiffy pellet ingredients in one place as your seed sprouts. Said ingredients include:

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Peat is an organic medium that provides plenty of nutrients for sprouting seeds. It is made from sphagnum peat moss, which has a high water-holding capacity as a result of its fluffy and light texture. Peat provides excellent root aeration, making it easy for roots to grow without interference.

Lime, Ammonium, and Fertilizer

Jiffy pellets are primarily made from peat moss. However, peat moss on its own is not ideal for sprouting seeds. That’s why they also include lime, which raises the pH of the medium. Jiffy Pellets have a pH of 5.5, compared to peat moss’s 4.4.

Jiffy Pellets also include fertilizer that provides seedlings with all their required nutrients for the first 1-2 weeks. You won’t need to add anything else as your seed sprouts. After transplanting, you can apply fertilizer, if desired.

How Big are Jiffy Pellets?

Jiffy pellets come in diameters of 24 mm, 33 mm, 41 mm, and 70 mm. The sizes vary to cater to the various needs of different seedlings. The larger Jiffy Pellet sizes are convenient to handle and are ideal for marijuana.

How to Germinate With Jiffy

Jiffy pellet instructions are easy to follow and will take you less than 20 minutes to set up, ensuring you’ll be able to start your cannabis seed germination project quickly and easily without wasted time or effort.

Choose Your Container

The first step is to place your pellets in a container. Ideally, you can use a Jiffy pellet tray. Ensure they do not overlap and leave a small border for water retention.

Water Your Jiffy Pellets

You can use tap water or distilled water. Keep in mind, tap water may have chlorine, which is harmful to your plant and can cause it to dry out faster. However, you don’t have to buy distilled water to prevent chlorine problems. Evaporate the chlorine by pouring your water, and let it rest for some time. Use very warm water, around 95oF, to help the pellet expand faster.

Plant Your Weed Seeds in the Jiffy Pellets

Place the seed in the middle of the enlarged pellet. Don’t place the seed too deep, or it will not sprout.

Cover Your Seeds

To avoid moisture loss, use a plastic cover – this usually comes with the pellets. You can also use plastic wrap. If you don’t have a Jiffy pellet dome, you will need to water again after around seven days. Avoid dropping water from the top; otherwise, you will disturb the developing seeds.

Transplant Your Plant into Some Soil

The seeds are usually ready for transplanting after two weeks. To do this, simply make a hole in the soil and place your pellet directly in that spot.

An Alternative to Jiffy Pellets to Consider

Coir is a popular alternative to Jiffy pellets , with Coconut Coir leading the trend. This material is made from coconut husks. If you choose this medium, use a coir Jiffy Pellet tray to hold it. Coir has excellent water retention capacities similar to peat, and it also facilitates the healthy development of roots.

Coir also repels insects and is reusable , so it can help you if you choose to expand your growing operations to include more cannabis plants.

What to do After Germination

After your seeds have germinated, they need to go into a pellet so the roots can spread out and grow. You may be wondering just how far in the jiffy pellet the seeds should be planted once it has germinated. You’ll want to drop the seed about a half-inch to an inch. If the seed has not sprouted, it’s okay to place it in the jiffy pellet – as long as it has soaked for at least 12 hours, but no more than 24 hours. If it has sprouted, drop the white root downward, so that the head of the seed is about an inch below the surface.

Damping-Off With Hydrogen Peroxide

Once your seed becomes a tiny plant, it’s now known as a seedling. This is great, except that seedlings are very delicate. You’ll want to be on the lookout for the number one seedling killer: damping off. Damping-off occurs when something (such as fungi or other diseases) causes the nutrients in your plant to stop being transported properly. The stem becomes mushy and falls over, and your plants will die young. The risk increases when there is minimal direct sunlight and air circulation, combined with warm temperatures and high humidity.

Eventually, your plant will be able to protect itself, but until then hydrogen peroxide is an excellent solution to this problem before it can begin. Hydrogen peroxide provides oxygen to the soil and reduces the chances of fungus spores developing.

To effectively prevent damping off, combine 1 tsp. of hydrogen peroxide with 2 cups of water. Use this solution to water the plants. Or you can soak the seed in 1% hydrogen peroxide before planting.

Frequently Asked Questions: Marijuana Seed Germination & Jiffy Pellets

Before signing off with this article, let’s take a few more moments to cover some key points included in this article to ensure you understand the basics of everything you’ll need to know when advancing forward in the seed germination process. We’ll also cover the answers to a few common questions new home-growers may want to address before getting started.

Germination is the process of seeds developing into new plants

All weed seeds need water, oxygen, and proper temperature in order to germinate.

It takes 1-2 weeks for weed seeds to germinate

We love jiffy pellets. They mimic the plant’s favorite natural medium; good soil.

The Bottom Line: What a Pot for Pot Can do For You

Germination can be tricky, but not if you have the right tools. The quality seedling starter kit from a Pot for Pot makes going from seed to seedling fairly effortless. It’s included in all our kits, or you can purchase it on its own. Simply select the seeds and provide some love and we’ll provide everything you need to grow healthy plants.

Do you enjoy an occasional marijuana smoke break but want to avoid costly trips to the local dispensary every time your bud stash starts to run low? Consider growing your own marijuana from the comfort of your home with one of our top-quality cannabis growing kits.

Learn how to cultivate marijuana. Find the best marijuana growing advice for seed germination, cloning, pruning, trimming, and curing to help maximize your yields.