Category Archives: Interspecific hybrids

All articles we have written about interspecific Jatropha curcas hybrids. Interspecific hybrids are crosses between different species within the same genus, for example Jatropha curcas x Jatropha integrrima. This type of breeding is considered difficult but has the potential to greatly enhance the range of genetic variance within a breeding population which again is a prerequisite for the development of elite cultivars of any species.

Large diversity in F2 interspecific hybrids not unexpected

Jatropha interspecific hybrids F2 generation available

Fruit on an interspecific hybrid

Fruit on an interspecific F1 hybrid

In the second half of August we successfully germinated the first three F2 generation seedlings from one of our interspecific hybrids. From now on we have fast growing F2 Jatropha interspecific hybrids for further breeding. While some research reports indicate that this type of interspecific hybrid is sterile or not interesting for breeding, we observe ongoing successful seed development on several of our early F1 plants deriving from different Jatropha curcas female parents. So we expect more additions to this F2 generation population in the near future.

As expected from applying simple Mendelian rules the F2 generation phenotypically exhibits much more diversity than the F1 generation.

All of the 3 seedlings we have so far appear strong and healthy. However, only the one on the left hand side seems to be as fast developing as expected while the other 2 are only moving along slowly.

Large diversity in F2 interspecific hybrids not unexpected

F2 seedlings of interspecific JcL hybrids 1 month old showing wide variation

We expect our F2 interspecific population to grow very fast over the coming months involving many different F1 interspecific parents with genetically very different Jatropha curcas parents in turn. Currently many F1s are initiating their first flowering. On the interspecific route of our breeding program this is really what we have worked on for almost 3 years. We have clearly begun moving into terrain which has not previously discussed in the scientific literature to the best of our knowledge.

This development marks another success in achieving our initial target to bring as much variability into the germplasm available to our Jatropha breeding platform for future improvements through selection. We are now able to start work on a real ground breaking next step we had been planning for since we did the first interspecific cross almost 2 years ago. With a little luck first results will become visible before the end of the year.

Further observations on the interspecific breeding route will be published here as much as possible, but of course we will have to start protecting our intellectual property where we reach the forefront of commercial hybrid development as we are expecting a meaningful patent application before too long.

Jatropha seed comparison

What makes an elite Jatropha cultivar

What exactly makes a certain Jatropha accession an elite cultivar? You can find the term everywhere but a clear definition is usually not given. Therefore I want to explore this subject today and attempt to demonstrate how an answer translates into  the daily work of a Jatropha breeder.

The term itself is simple: elite means something like “better than average” and cultivar is just an abbreviation of “cultivated variety”. But what does it really mean to us?

The most general and holistic objective when breeding for an improved commercial crop is of course always maximum profitability for the farmer. As this is self explanatory only a next level break down of objectives can offer some insight. In our program we use 4 groups of traits which in turn again break down into the real, identifiable traits when observing a selected plant.

The groups we use at Bionic Palm are:

  1. Seed Quality
  2. Seed Yield
  3. Agronomy
  4. Pest resistance.

We are looking for improved traits in all of the above groups. Let’s have a quick look at each group to see some of the aspects a breeder is really looking for every day.

Seed Quality
For us this includes several individual traits, all linking back to profitability of the crop of course.

  • Toxicity
  • Uniformity within the seeds
  • General healthiness of the seeds
  • Chemical composition of oil and cake.

No further explanations needed here I think.

Seed Yield
This is the most relevant for most people discussing Jatropha, but what are we really talking about, when we use the term? 

Many people would just want to count the seeds or go by tons per ha. An approach that would translate into something like number of seeds harvested per plant per year. This again translates into bottom line traits like branching (self or pruning induced), number of fruit clusters or fluorescences per branch, number of fruits per cluster (= number of total flowers in a fluorescence plus male:female flower ratio). Not too difficult to understand, once you know the Jatropha exclusively flowers at the top of a branch.

However, that’s not even half the truth. In our understanding the high level measurement must actually be average amount of crude oil obtained per ha (or acre) combined with average amount of cake per ha.

We have recently taken a closer look at seeds obtained from the various hybrids we produce. A comparison shows significant differences in seed weight and oil content which are actually quite stunning. And the spread is increasing the longer we breed…

Jatropha seed comparison

The typical measurement 100 seed weight varies from around 45g to 98g in our current accessions and oil content ranges from 25% to clearly over 50%. That means with an equal number of seeds per ha I can actually have up to 4 times the oil!

Agronomy

 Lets move on to traits important for agronomic reasons. Many of them are related to the cost involved in farming Jatropha and the most to the ease of harvesting.

There is the uniformity of fruit ripening which determines how many times a picker has to pass a plant per season, then there is canopy structure and size of fruit clusters which determine the ease and speed of harvesting. Other important agronomic traits are self branching (without the need of pruning). Also drought resistance or rather the reaction to water stress is an important trait. We find that there are tremendous differences between accessions. The difference is at what level of water stress a plant stops female flowers, no flowers at all, throws off developing fruits and more. We see a clear pattern of those reactions differing greatly among accessions. I also have to admit that so far the best performers are the weakest in terms of water stress resistance.

Pest Resistance

Many of the common publications about Jatropha suggest that their toxicity would protect them from pests. This is not the case! The most serious pest we are confronted with is the papaya mealy bug. It comes and goes with water stress. There is no correlation with toxicity at all, however a clear correlation with resistance against water stress. And it should be at least mentioned here that the available phosphorus in the soil plays a major role in any plants ability to handle water stress. Therefore the right soil amendments like mycorrhiza can make a huge difference in addition to the best suited accessions if you plan to plant Jatropha in the desert. :-)

I am aware that this review of relevant traits in Jatropha is anything but exhaustive, but I don’t want to bore my readers too much with the obvious. Therefore I tried to point out only a few of the less discussed but important aspects of this fundamental subject on the way to true Jatropha elite cultivars.

The role of heterosis in Jatropha breeding

Today I want to take a closer look at the relevance of heterosis in Jatropha breeding. Heterosis or hybrid vigor, or outbreeding enhancement is “the increase in growth, size, fecundity, function, yield, or other characters in hybrids over those of the parents”. Find more information on heterosis on Wikipedia.

Heterosis effect can be forecasted based on this genetic variability map

Almost 3 years ago we started all our breeding efforts from this genetic variability map created by our friend and advisor Tommaso Barbi working at Geneticlab in Trieste, Italy at the time.

The parental accession base of the Bionic Palm Jatropha breeding program is not a 100% overlap, but we do have quite a significant number of key accessions in our program which are also shown in the map while many others can be linked to one of the clusters genetically or phenotypically.

Simply speaking, crossing more distant accessions from the map increases the likelyhood of significant variability in the hybrids genetically expressing itself in a higher degree of heterozygosity. Another crazy insider term from molecular biology, I know. Heterozygosity describes the amount of differences between corresponding alleles in a DNA sequence. Homozygosity is the term for the opposite situation of corresponding alleles being similar.

It has been found that heterozygosity usually correlates with strong heterosis effects. In the map above a cross between more distant accessions leads to a higher heterozygosity in the resulting hybrids and therefore higher levels of heterosis can be expected. Only about 6 months ago we started to be able to observe different heterosis levels in our hybrids.

The molecular basis of heterosis is still not fully understood. Current believe among scientists is that healthy alleles across numerous genes from one parent compensate for dysfunctional alleles at those genes from the other, distantly related parent (and vice versa for a second set of genes). A plant exhibiting a high level of heterosis would thus be the genetically most healthy.

This explanation fully corresponds with the opposite effect of inbreeding depression. Here parts of the parental alleles become dysfunctional or silenced because there is too little genetic difference between the two parents. The very strong expression of heterosis we observe when crossing a semi-commercial Jatropha accession from Africa or Asia with a wild accession from the center of origin therefore suggests at least a mild inbreeding depression on a very narrow genetic variability in the germplasm originally introduced by the Portuguese seafarers when they first distributed Jatropha around the world.

So far observations are limited to leave size and, at a smaller extent, stem diameter. Over the coming month fruit and seed sizes will become visible too. With the leaves we quite commonly measure lengths of up to 20cm where the usual average is no more than 10cm.

The following picture demonstrates the phenomenon very clearly.

Heterosis in Jatropha breeding can be observed in this intraspecific hybrid plant

The small plant in the front germinated in November 2011 and is a cross of 2 accessions with distinct phenotypic differences but, genetically very similar, they are both from the upper left cluster in the map. The much larger plant behind in the center germinated in March 2012, 5 months later, and represents a cross between the upper and lower left hand clusters. Average leave length is almost double that of the plant in the front and of course the overall size is significantly different.

 A heterosis effect is also visible in many of our interspecific hybrids, but notably less so in those which have a very short seed-to-seed time of less than six months. A behavior which can be attributed to negative heterosis (=shorter seed-to-seed cycle). Interspecific heterosis can be observed in the following picture.

Heterosis in Jatropha breeding

The plant on the far left hand side shows by far the most dramatic heterosis effects. Followed by the next 2 plants to the right with heterosis also clearly noticeable. All 3 happen to be non-toxic interspecific hybrids. Then we see an early flowering and fruiting interspecific hybrid exhibiting no visible heterosis. Similarly at last, on the far right some regular Jatropha can be seen. From left to right we go from high level heterosis to none at all.

In front a rare chili pepper can be seen which is currently tested for drought resistance.

The findings described in this post have been highly unexpected. We are currently waiting for the first seeds from plants exhibiting strong heterosis to mature so we can check seed weight and oil content. Based on the results we will have to decide if we are going to open up a new heterosis breeding route in addition to our current approaches.

Early BC1 collection

Jatropha hybrid backcrosses available for further breeding

Before our proof-of-concept phase officially came to an end on June 30th, 2012 we succeeded in taking our efforts to the next level. Our first Jatropha hybrid backcrosses have germinated and reached seedling stage by mid June. This achievement marks an enormously important step forward for the Bionic Palm Jatropha breeding program. 

Jatropha hybrid backcrosses exhibit enormous genetic variability even at the seedling stage

The above picture shows the huge variability among our earliest BC1 seedlings. Over the coming months this collection will grow into the hundreds. We aim for a BC1 population incorporating all of our original parental germplasm and most of its cross combinations. Thus we will gain the maximum possible genetic variability base which we can later draw from when we breed for truly superior commercially usable hybrids.

What we define as our BC1 population is the result of back-crossing interspecific Jatropha curcas x Jatropha integerima hybrids into our parental Jatropha curcas accessions.  This is a standard breeder’s approach if he wants to transfer specific traits (certain properties) from a related species into his species of interest. However, we are not into pedigree breeding at this time. Our primary aim is still to increase genetic variability of our population to the utmost possible.

We are looking for the greatest possible heterozygosity in these plants which can be assumed to be 100% in many cases. According to scientific research reports on the genetics of Jatropha this can only be achieved through an interspecific breeding route. While this kind of approach is very time consuming we do expect to gain a substantial shift in the Jatropha genetics.

Interspecific hybrid H1.2

Jatropha interspecific hybrids doing well

Today we take a quick look at one of our oldest Jatropha interspecific hybrids. It’s less than 7 months old and about 1.5 m high. Over the last 3 months it has been flowering almost constantly. Fruit development is becoming more and more robust, so we expect larger quantities of healthy seeds in the near future.

As this type of hybrid represents an intermediate stage in one of our originally charted out breeding paths male and female flowers will be used for further breeding. However, in order to exploit the full genetic variability of such a unique hybrid a next generation direct offspring (F2) will be of great interest.

Only 3 days ago it was transplanted to a large decorative pot and placed at a prominent spot in front of the house because this will be the most memorable plant of the BPL Jatropha breeding program for a long time to come.

BPL H1.2 interspecific hybrid

After intense rainfall once again we see a large number of fruits developing. While we already harvested an early seed we had no germination success yet. Difficulties in fruit development actually seem to be a trait inherited from the pollen donating parent. However, we are more than confident to break that barrier in the near future as we have broken the barrier before on the male parent after some experimenting.

Posted – for the first time – mobile from WordPress for Android on my good old Galaxy Tab

Next milestones on the way forward

A couple of weeks ago we have successfully reached a very important milestone of our breeding program: the first interspecific hybrid was flowering. As of today we are in the middle of a sister hybrid flowering with at least 4 others approaching the same stage. Also, the initial F1 plant is already showing the next 2 inflorescenses which might indicate a very active flowering trait.

So what have we done since the F1 flowering commenced and what are the next steps going to come?

First of all on the very first pollinated hybrid we have 5 fruits developing well. They have all been manually pollinated with various non-toxic pollen donors over 3 weeks ago. In addition, this hybrid provided pollen to a most promising non-tox variety luckily flowering at exactly the right time. So we have 3 BC1 (first generation back cross) fruits developing.

The second hybrid which is in the middle of an extensive multi-week flowering processes right now has opened 13 female flowers so far over the last 5 days. They have all been manually pollinated utilizing 4 different pollen sources. This flower shows a lot more buds that are going to open more female and a lot of male flowers over the coming days. The very delayed process of individual flower openings is extremely helpful (obviously a trait crossed into the JcL from the other Jatropha species used), as it increases the chances of many interesting successful cross pollinations.

For this type of interspecific F1 hybrid we already have over 50 samples deriving from 8 genetically different JcL accessions. This type of crossing will become a standard process step with all new JcL accessions we aquire. Currently we have over 25 proven distinct accessions growing with initial flowering expected between 1 and 10 months out. About 1/3 tested as non- or low-toxic.

Some of the readers might have by now identified the other Jatropha species used in this hybrid. Actually this route of hybridization has been described in scientific reports several times in the recent past. The more we are surprised how little information is publicly available about the characteristics and various crossing options it offers. It seems, that so far nobody explored this hybrid in-depth instead of rushing on to the next generations. Seeing the huge potential I find this a bit astonishing. It seems to be the result of many research projects being tied to limited time and funding. As it takes 18-24 months to get a flowering F1 hybrid and most public research funding today is on a 3 year basis, we can understand, how the project time remaining was running out. Therefore, we might very well be the first breeding program that has the capability of scratching a little deeper into the vast potential variability of that F1 generation. This will of course quickly lead to hundreds or even thousands of plants in the first generation alone and result in millions of plants one or two generations further down the road. So we are already looking for ways to reduce those enormous numbers.

Flowering H1 hybrid

Interspecific Jatropha hybrid flowering

Flowering H1 hybridWe have arrived at and actually already passed one of the most important milestones of our breeding program: our first interspecific  Jatropha hybrid has opened flowers this morning.

While we are still awaiting the outcome of a large number of other breeding results, this hybrid clearly exhibits several of the expected traits: a very short period of time from seeding to flowering (less than 4 months), a very positive female:male flower ratio and a relatively good resistance against the mealy bug.

This plant is a key outcome of our proof-of-concept phase and represents a highly successful intermediate step on the way to our core breeding objective: the creation of a high yielding, non-toxic Jatropha variety exhibiting a high level of horizontal resistance specifically in West African ecosystems.

The three visible female flowers have already been pollinated manually according to the pre planned breeding paths of which the most important one will be back crosses with selected non-toxic and toxic Jatropha accessions with superior traits. According to current experience we expect a next generation of breeding plants within 6 months.

 

The Bionic Jatropha breeding program in 2012

Since we started our JcL breeding program we focused primarily on elite non-toxic hybrid cultivars. We have completed the main phase of a proof of concept (PoC) meant to significantly reduce the commercial risk involved in a full scale program go ahead.

By the end of 2011 we have succeeded in showing the validity of all three of our principal approaches, one involving traditional intra specific hybridization, one an already well documented inter specific hybridization path and one a completely innovative bottom up inter specific hybridization method.

We have recently decided, to add a 6 months extension to the PoC to gather more useful planning data especially by reaching the third generation of hybrid breeding, but also by further verifying and confirming protocols developed over the last 18 months.

During the PoC we have worked with 5 different toxic JcL accessions from Africa and Asia as well as 4 distinct non-toxic accessions from the center of origin and a number of accessions from 2 related species. All that parental material has been used to establish a stable, mature selection of germplasm ready to be used in larger scale breeding approaches.

We have also expanded our carefully selected population of parental material for further breeding by another 8 unique non-toxic and 15 toxic JcL accessions. Especially useful is a male sterile, high yielding non-toxic accession and a line exhibiting natural polyploidy.

Currently we have approximately 50 intra and interspecific hybrids growing and we expect some of them to set flowers within the next few weeks. With the resulting BC1F1 and F2 generations we will soon enter the core breeding phase.

Over the coming 6 months we will initiate some systematic feed trials with the first generation non-toxic material we have planted at our farm. For this purpose we have established a small poultry test farm comprising of chicken, guinea fowls and turkeys.

 

Breeding for resistance

A major reason that motivated us to look into JcL breeding goes back to a devastating pest attack. Starting about 2 years ago the Papaya Mealy Bug arrived. It settled down heavily on all our JcL hedges and also infected tomato and pepper plantings. During the very dry year 2010 we lost almost all our crops to this bug.

Studying the little options available to fight the plague (of course the industry is willing to sell you tons of different pesticide products, but they don’t really work) of course breeding for resistance was an interesting way forward. As we had started looking at breeding for yield and removal of negative toxic effects anyway, it soon became clear what the three primary objectives for our breeding program would be: (1) yield (2) absence of toxicity and (3) resistance.

Breeding for resistance is an extremely controversial subject and past attempts have done a lot of damage to specific crops and ecosystems that play a major role in today’s overzealous use of chemicals in agriculture.

Recently I stumbled over the very enlightening books of Raoul A. Robinson, especially the one called “Return to Resistance”. (All of Mr. Robinson’s books can be downloaded for free from the internet, just google his name) In this book he lays out the problems around vertical (one gene) resistance and makes the case for horizontal resistance, with the latter being a potential remedy for many of the current food shortages.

Horizontal resistance is a multi-gen trait of a plant that is permanent similar to the kind of resistance wild plants obviously have. In commercial corps in can be achieved through continuous cycles of mass selection from self crossing accessions which will inevitably improve resistance more and more until an upper limit is reached. The larger the range of genetic variability in the breeding population, the better.

We have decided, that we will give this approach a similar chance as the “Mendelian” approach where we cross breed intra- and interspecific hybrids looking for the best combination of traits (pedigree breeding). We actually believe, that introducing interspecific hybrids with their greater genetic variance can help improve the start-up population for the mass selection population.

Anyone with some interest in the subject should definitely have a look at Raoul Robinson’s books. You don’t have to be an agronomist or a plant breeder to understand his work. I can only highly recommend it.