Construction and characterization of a partial binary bacterial artificial chromosome (BIBAC) of Agave tequilana var. azul (2X) and its application for gene identification

The structure and organization of the genome of Agave is still unknown. To provide a genomic tool for searching sequences of the genus, we built and characterized a binary (BIBAC2) genomic library of Agave tequilana  Weber var. azul. Clones of the library had an average insert size of 170 Kb. The frequency of inserts with  internal Not I sites was 30% and only 5% of the library showed organelle contamination. The library was  assessed using probes with high homology to repeated regions (retroelements and rDNA regions), genes  involved in the resistance to diseases (NBS-LRR) and genes related to late embryogenesis (LEA).  Recombinant clones that hybridized with each of the probes were identified. Our results indicate that the  obtained genomic library is suitable for the identification of sequences of interest, for genetic mapping and for studies of gene regulation and expression.Key words: Binary bacterial artificial chromosome (BIBAC), Agave tequilana, genome, clones

Genetic characterization by amplified fragment length polymorphism (AFLP) markers and morphochemical traits of Carica papaya L. genotypes

Carica papaya L. is a native fruit from Central America and Mexico and it is an economically important fruit. As a pre-breeding genetic study, the variability of both parents (L7 and M22) and the F1 individuals derived from their crosses (L7 × M22), was evaluated in terms of 32 morphochemical traits, and contrasted with their genetic diversity indicated by amplified fragment length polymorphism (AFLP) markers. According to morphochemical traits, L7 and M22 were grouped in two different clades. The first group included L7 and 13 genotypes from the F1, while a second group included the parent M22 and 15 other genotypes from the F1 progeny. The analysis based on morphochemical traits showed an average correlation of 0.652 among genotypes. For AFLP analysis the combination of the primers E-ACA/M-CTA had the best polymorphic index (72.73%). When they were grouped based on AFLPs markers, it was confirmed that both parents are genetically distant, and they were again grouped in two different clades. Five genotypes from the F1 population were grouped in the same clade as L7 and shared 55% similarity. Twenty six genotypes were grouped in the same clade as M22, showing 63.3% similarity. Another 12 genotypes (mainly female genotypes) were grouped in a third independent clade. This relative general agreement between the grouping based on a large number of morphochemical traits (including both plant and fruit traits) and that based on its genetic diversity using AFLPs, suggests that morphochemical characterization, together with genetic analysis by AFLPs, can be complementary and useful techniques for the identification and assessment of genetic diversity within C. papaya L. genotypes, that should be useful for genetic breeding programs of this important species.Key words: Morphological markers, AFLP markers, genetic similarity, Carica papaya L

Diversidad Génetica de Agave angustifolia Haw. Silvestre y cultivado de Oaxaca, México

Instituto Politécnico Nacional CIIDIR Oaxac

A quantitative PCR approach for determining the ribosomal DNA copy number in the genome of Agave tequila Weber

Background: Agave tequilana has a great economic importance in Mexico in order to produce alcoholic beverages and bioenergy. However, in this species the structure and organization of the rDNAs in the genome are limited, and it represents an obstacle both in their genetic research and improvement as well. rDNA copy number variations per eukaryotic genome have been considered as a source of genetic rearrangements. In this study, the copy number of 18S and 5S rDNAs in the A. tequilana genome was estimated, and an absolute quantitative qPCR assay and genome size was used. In addition, an association between the rDNAs copy number and physical mapping was performed to confirm our results. Results: The analysis were successfully applied to determine copy number of 18S and 5S rDNAs in A. tequilana genome, showing high reproducibility with coefficient of variation (CV) values of 0.014–0.0129%, respectively. A variation of 51 times in the copy number the 18s regarding 5s rDNA was found, thus contributing to genome size of 1.47 and 8.38 × 10-3%, respectively. Similarly, data show a linear relationship (R [2] = 0.992) between rDNA copy number and the detected signals for each of the loci by FISH. The comparison of the rDNA copy number of agave showed differential relationship with other organisms and it may be due to evolutionary ecology. Conclusions: Results show that the proposed method a) can correctly detect the rDNA copy number, b) could be used as species-specific markers and c) might help in understanding the genetic diversity, genome organization and evolution of this species

Sea turtle hatchling carapace as a source of high quantity and quality DNA

Hatchling DNA provides valuable information on sea turtles. Samples can be obtained from dead hatchlings or embryo, or, when live animal samples are needed, from blood, flipper or carapace. We compared 120 DNA extractions from flipper and carapace tissue of dead and live hatchlings. There were significant differences in DNA yields from the different tissues, but no significant differences in DNA purity. Some flipper samples yielded low amounts of DNA, while the carapace tissue consistently produced high yields of good quality DNA. This suggests that carapace represents the best option for tissue sampling of hatchling sea turtles in genetic research

Data from: Genetic variation, multiple paternity and measures of reproductive success in the critically endangered hawksbill turtle (Eretmochelys imbricata)

The Yucatán Peninsula in Mexico contains some of the largest breeding groups of the globally distributed and critically endangered hawksbill turtle (Eretmochelys imbricata). An improved understanding of the breeding system of this species and how its genetic variation is structured among nesting areas is required before the threats to its survival can be properly evaluated. Here, we genotype 1195 hatchlings and 41 nesting females at 12 microsatellite loci to assess levels of multiple paternity, genetic variation and whether individual levels of homozygosity are associated with reproductive success. Of the 50 clutches analyzed, only 6% have multiple paternity. The distribution of pairwise relatedness among nesting localities (rookeries) was not random with elevated within-rookery relatedness, and declining relatedness with geographic distance indicating some natal philopatry. Although there was no strong evidence that particular rookeries had lost allelic variation via drift, younger turtles had significantly lower levels of genetic variation than older turtles, suggesting some loss of genetic variation. At present there is no indication that levels of genetic variation are associated with measures of reproductive success such as clutch size, hatching success, and frequency of infertile eggs

Data from: Genetic variation, multiple paternity and measures of reproductive success in the critically endangered hawksbill turtle (Eretmochelys imbricata)

The Yucatán Peninsula in Mexico contains some of the largest breeding groups of the globally distributed and critically endangered hawksbill turtle (Eretmochelys imbricata). An improved understanding of the breeding system of this species and how its genetic variation is structured among nesting areas is required before the threats to its survival can be properly evaluated. Here, we genotype 1195 hatchlings and 41 nesting females at 12 microsatellite loci to assess levels of multiple paternity, genetic variation and whether individual levels of homozygosity are associated with reproductive success. Of the 50 clutches analyzed, only 6% have multiple paternity. The distribution of pairwise relatedness among nesting localities (rookeries) was not random with elevated within-rookery relatedness, and declining relatedness with geographic distance indicating some natal philopatry. Although there was no strong evidence that particular rookeries had lost allelic variation via drift, younger turtles had significantly lower levels of genetic variation than older turtles, suggesting some loss of genetic variation. At present there is no indication that levels of genetic variation are associated with measures of reproductive success such as clutch size, hatching success, and frequency of infertile eggs

Ei-genotypes

The database contains the genotypes of 41 nesting hawksbills and 1,188 hatchling that were successfully genotyped at 12 polymorphic microsatellite loci. Paternal reconstructed genotypes are also includes. The name of the rookery is abreviated where: XV=Xicalango-Victoria, CK=Chenkan, CE=Celestun, CL=Las Coloradas, CY=El Cuyo, H=Holbox. Id letters identifies each nesting females, her offspring and the male that sired that clutch. In those cases where offspring contain and Id letter that is not on the females or fathers id indicates that it is a successive clutches of the same previously reported females (i.e. if there are offspring with and Id=B, but there are no females with these Id, then is a successive clutch from female A). In the cases where males Id have letter and number indicate that those multiple males sired a same clutch indicating multiple paternit

Caracterización de accesiones de papaya (Carica papaya L.) a través de marcadores AFLP en Cuba

Los marcadores moleculares son herramientas valiosas en los estudios genéticos en plantas, y están siendo empleados exitosamente en programas de mejoramiento principalmente en la elección de progenitores y en la selección. El polimorfismo observado mediante la técnica molecular AFLP (Amplified Fragment Length Polymorphism) ha sido de utilidad para estudios de diversidad genética en frutales. En el presente trabajo se realizó la caracterización molecular de 12 accesiones de papaya (Carica papaya L.) del banco de germoplasma del Instituto de Investigaciones en Fruticultura Tropical (IIFT), empleando la técnica AFLP. Se evaluaron seis combinaciones de iniciadores para la amplificación selectiva, las cuales amplificaron un total de 431 bandas con 73,3% de polimorfismo. El número total de patrones de bandas identificados fue igual en todas las combinaciones utilizadas, con un porcentaje de identificación alto, lo que sugiere que dichas combinaciones pudieran ser empleadas para estudios de variabilidad genética en papaya. En general, los resultados presentados demuestran que existe diversidad genética entre las accesiones evaluadas, lo cual constituye un reflejo del origen que presentan los genotipos analizados a partir de la introducción de materiales foráneos y la polinización abierta de un grupo de materiales selectos. Por tanto, se recomienda retomar la prospección y selección de accesiones locales, así como la introducción de nuevos genotipos foráneos, como dos vías fundamentales para aumentar la diversidad genética presente en el banco de germoplasma de papaya de Cuba. Palabras clave: marcadores moleculares, polimorfismo, diversidad genética

Reproductive success

Information of nesting females and reproductive success data of their clutches. Females that were marked were considered as neophytes or first time nesters, while recaptures were considered as experienced or older females. CCL= curve carapace lengt