木薯作物遗传资源、生物技术与育种

Luiz JCB Carvalho 陈松笔

摘  要：木薯貯藏根具有提升世界热区人类食物营养品质的特性。本文综述了导致木薯（Manihot esculenta Crantz）贮藏根特性发生显著变化的变异、进化、遗传资源和驯化相关的中性遗传论题。传统育种通过更新资源信息，包括遗传资源、遗传多样性、基因组学、转录组学、蛋白质组学和代谢的方法改善木薯贮藏根特性，并且介绍了木薯贮藏根淀粉含量、蛋白质含量、类胡萝卜素含量和常规品种的改良。

关键词：木薯;贮藏根;营养品质;遗传资源;遗传多样性;传统育种

中图分类号：S533         文献标识码：A

1  Introduction

Cassava （Manihot esculenta Crantz）， belonging to Euphorbiaceae， is currently ranked as the sixth most important crop produced， and is a staple food for almost one billion people in the world[1]. It is the major source of calories in sub-Saharan Africa， where it is grown primarily for its starchy storage roots[2]. Cassava is an inexpensive source of starch and is currently being developed for industrial uses as well as a source of animal feed， primarily in Asia. Nonetheless， most of cassava in the world is consumed by subsistence farmers in Africa and Latin America[2]. Many basic biological traits of the crop and its closely related species have gone understudied until recently， in stark contrast to the extensive

work on cash crops such as corn， wheat， rice， and soybeans. The spoilage of cassava storage roots occurs quickly after harvest through physiological deterioration caused initially by oxidation followed by colonization and growth of various saprophytic microorganisms. In addition， cassava plants are susceptible to various pathogens， including a bacterial blight and the cassava mosaic virus， a devastating pathogen that can reduce crop yields by as much as 70%. The challenge for crop breeders is to increase nutritious quality of cassava storage roots （CSR） and pathogen resistance plants， while reducing postharvest physiological deterioration （PPD）. Cassava， as a cash crop is low because of its starch， quality， making it less desirable as a source of raw material for the food industry. Any modification of cassava to enhance its value as a market crop would have direct and positive effects on the lives of families. As it is discussed in this review， landraces of cassava that sequester carbohydrates other than amylose have great potential as cash crops. Traditionally， many plant breeders have followed to landraces， wild ancestors， and closely related species as a source of traits for futuro crop improvement[3]. We have investigated the wild ancestor of cassava， identified its putative origin site of domestication， and are studying the relationship of the cassava crop， and characterizing the biodiversity of landraces in the Amazon River Basin in Brazil. This review reports and describes the potential reservoirs of germplasms for the improvement of cassava crop in Brazil， and attempts to show that cassava serves as a model system for understanding evolution in a recently arisen plant genus and for understanding domestication in a perennial， clonally reproducing crop that uses an underground vegetative storage organ as food consumed by humans[3-9].

2  Cassava species

The genus Manihot （Euphorbiaceae） shows about 98 Neotropical species with plants range in habit from herbs to small trees that are distributed in two centers of origin documented[10-26]. One locates in Brazil with 80 species and another in Mexico with 17 species which is updated， including Brazil M. esculenta Crantz distribution from East/West along the South Amazon boarder as summarized in Fig. 1.

3  Geographical Manihot species distribu- tion in Brazil and their evolution

Manihot species distribution in Brazil and their phylogenetic relationships to Neotropics species have been recorded using geographical information， morphological traits and molecular markers. This review summarized and updated as shown in Fig. 1 to understand the genetic structure and conservation[25] and illustrated in Fig. 1 that shows a compiled and updated recorded analyzed information.

4  Brazilian Manihot species relationships： a phylogenetic kinships study

Studies on Manihot species evolutionary relationships by Allem and others indicated a cassava crop gene pool composed of four closely related Manihot species （Fig. 1C）， including M. flabellifolia ssp. flabellifolia， M. flabellifolia ssp. peruviana， M. flabellifolia ssp. esculenta， and M. pruinosa. These studies allow us， together with genomic information， to infers that M. flabellifolia ssp esculenta is indeed derived directly from M. flabellifolia ssp. flabellifolia as compiled recorded evidences updated information thought-out Fig. 1 using references[20-28].

5  Agro-biodiversity of cassava crop in the center of origin and domestication in Brazil： storage root， a vegetative storage organ for food

Owing to poor preservation of organic remains in humid environments， direct evidence of early manioc （M. esculenta Crantz） cultivation is exceptionally rare in datable archaeological contexts. Above our research summarized here and offer new insights into spatio-temporal framework of the initial domestication and early spread of manioc in the Neotropics. Integrating evidence from comparative plant genetics and paleoethnobotanic starch analysis to contribute to the archaeology of manioc origins， this review finds that （1） the strongest candidate for the botanical origin of domesticated manioc the wild progenitor of the root crop is the species M. esculenta ssp. flabellifolia; （2） the geographical origin of manioc the bionic in which the progenitor evolved is most likely in the savannas， the Brazilian Cerrado， to the south of the Amazon rainforest; （3） the Cerrado is also， in our best estimate， the region of agricultural origin of initial cultivation; （4） domesticated manioc had spread from the agricultural origin by the early Holocene， possibly as early as 10 000 years ago， but certainly by 7000 B.C.; and （5） domesticated manioc was a readily available plant in most habitats of the Neotropics by the mid-Holocene， at least 6500 years ago （Fig. 1）. Evidences on the cassava domestication in the lower Amazon river base region as recorded above， one expects that this would be a geographic area where humans have had a long traditional association with cassava （Fig. 2）. Many visitors to the region have noted the diverse uses of cassava among villages. Cassava meal is used in sauces， flour is baked into flatbreads， the leaves are ground and cooked， and several fermented drinks are produced from the storage root as well as storage root pickles. This diversity of uses is quite different from the use of cassava cultivated in much of the rest of the world， where it is grown as a source of starch for flour. Moreover， in the Amazon cassava is grown mostly in small， intercropped fields or in backyard gardens that often contain several distinct landraces of cassava. This is in contrast to the monoculture of cassava observed for improved varieties grown in many parts of the world. Given that both the uses of cassava and the mode of cultivation are more diverse in the Amazon region there may be underlying variability in key agronomic characters for cassava that could be useful in addressing the challenges cassava faces as a crop. The discovery of the site of domestication allows agricultural biologists to focus their areas of study and collection. Several field trips to the Brazilian Amazon to learn of new uses and varieties of cassava and to collect diverse storage root variants have been performed. Smallholder farmers， isolated rural communities， local markets， and regions with different systems of cassava cultivation were visited in the states of Mato Grosso， Rondonia， Amazon， Para， Marajo Island， and Amapa. The landraces in these regions showed an astounding diversity in unusual storage root traits related to root shape， color， and structure as well as carbohydrate contents and types. A field test for starch based on iodine allows one to identify starch in a cross-section of the storage root and to identify the types of starch and the patterns of starch distribution. Cross-sections of various landraces of cassava clearly show diversity in both the presence or absence of starch and the pattern of starch distribution. Biochemical studies of the carbohydrates of these landraces revealed a new type

6  Genetic studies with natural populations of cassava （M. esculenta Crantz）

Morphological mutations affect the outwardly visible and measurable properties in cassava plants， such as shape， size， color， chemical composition and starch type. A germplasm collection of anomalous cassava plant has been organized and is maintained at the general cassava germplasm collection （Colbase） in Embrapa Genetic Resources and Biotechnology （Brasilia， DF. Brazil）. The Cassava Mutant Collection （CMC）， reported first time in this review as a gasp description of seven observed mutant plants （Fig. 3） with unusual morphological and biochemical characteristics， that includes 1-Dwarf plant， 2-Anomalous flower type （Hermaphrodite）， 3-Anomalous leaf petiole size， 4-Variegated leaf type， 5-High lycopene content storage root， 6-Sugary storage roots， 7-zigzag intermodal stem. The genetic diversity observed in germplasm collections are reported below and listed in Tab. 3[31-40].

7  Germplasm collections of cassava （M. esculenta Crantz） plants materials organi- zation， availability， and studies

Today the field of cassava plant genetics continues reliance to address many of the same questions about natural plant variation posed by early geneticists while integrating developments in genetic resources， molecular， genome and computational biology information. Here we explore natural variation in the genus Manihot focusing on how it evolved， how it was domesticated， how it is distributed in both domesticated and wild Manihot species， how it conditions complex phenotypes and how it can be efficiently utilized in modern cassava crop improvement to improve CSR nutritional qualities. We have pioneered studies demonstrating that because cassava was domesticated in the lower Amazon region， one expects that this would be a geographic area where humans have had a long traditional association with this plant， allowing the analytical studies as proposed at the references[41-44]. Several field trips to the Brazilian Amazon were made to learn of first new uses and local cassava plants to collect diverse storage root variants. The landraces in this region showed beyond belief diversity in unusual storage root traits related to root shape， color， and structure as well as carbohydrate content and type that may be underlying variability in key agronomic characters for cassava crop improvement. The cassava diversity has been taken by a collaborative initiative among Embrapa research units coordinated by Embrapa Genetic Resources and Biotechnology （Tab. 1） in order to explore the genetic basis of variation in cassava and its ancestors. Together， biological and informational resources are being rescued， organized and are presented under Manihot species portrayal， phenetics of Manihot species， their geographical distribution， domestication traits， general germplasm collection （Tab. 1） organization （Colbase）， regional work collection organization （Tab. 1）， breeding population collection （BPC）， and cassava mutant collection （CMC）. The genetic resources underline the foundation of continue studies on genomic， proteomic， breeding and utilization of cassava in Brazil and other parts of the world is found in the website linking below. To access diversity in cassava germplasm， experiments were performed under a license from the Genetic Heritage Management Council （CGEN） as required[40] and follow the approval studies from the local Ethical Review Panel of Embrapa Genetic Resources and Biotechnology[41].

Germplasm collections， as documented （Tab. 1）， were organized based botanical species isolations， and geographical genetic isolates as initial genetic identity and used under hypothetical apparent population evolutionary biology approach. TIME0 （Center of Origin & Domestication） refers to time zero， considering early genetic isolating events such as speciation genetic events including early domestication， mutations， recombination， drift selection of plant population in the center of origin and domestication of Manihot species， mainly M. flabellifolia ancestors in Amazon region （Brazil） as previously reported. TIME1 （Secondary Region） refers to cultivated forms after migration and volution-mutation/selection， lineage of domesticated cassava （M. esculenta Crantz） and traits variations under studies in local landraces， moderns cultivars in Brazil （landraces）， Secondary Regions， landraces， breeding clone populations， and Candidate Gene Genealogy refer to lineages that were defined base on carotenoid， starch biosynthesis， accumulation and molecular marker groupings as previous reports.

8  Secondary Regions （mutant plants， novel haplotypes）

Studies reported in Manihot species， and cassava （M. esculenta Crantz） plants traits variation， their possible genetic diversity at population level， and evolution were sampled to approach with genomic， transcriptome， proteome， metabolism， and conventional breeding. Experimental documentations are available in this review as shown at Fig. 1-10.

9  Genome sequencing studies

Progresses on knowledge about the cassava plant genome and proteome sequences are available in several public data bases as listed in Tab. 2.

10  Transcriptome studies

Transcriptome studies were undertaken with microarray data analysis of landrace CSR pigmented variation in relation to breed commercial line and ancestor[48-51]. These data allow us to speculation on gene regulatory functions unique for white CSR and single mutant in genes mutated in pink or intense yellow landrace.

Starch synthesis pathway： survival related genes. The hypothesis diagram showed in figure 4 provided information guideline of sampled population of plants to search for biochemical traits， variations， and candidate genes to understand carotenoid synthesis pathway as well as amount of free sugars （mainly glucose） starch and starch type. The proposed ancestor of cassava， M. flabellifolia， shows white storage root in nature. The variants of pigmented cassava storage roots and high protein contents have been reported[17]. Thus， white storage root is hypothesized to be the ancestral state of the genes related to these traits. The pigmented storage root phenotype is thought to have originated as a naturally occurring variant[17]. There is no known fitness advantage to store carotenoids in the storage root， and the current prevalence of intense yellow， pale yellow， and pink phenotypes is entirely the result of artificial selection that occurred during cassava domestication recently[15， 19-21] as well as sugary[16] and high proteins content associated with pigmented cassava storage roots.

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