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We report the accurate determination of the allelic configurations of a total of eight new citrus tetraploid hybrids by means of SSR analysis, coupled with capillary electrophoresis, and PCR based dosage effects. Tetraploid hybrids were spontaneously obtained from different interploid crosses (2x × 4x) between diploid ‘Femminello’ lemon and the allotetraploid somatic hybrid (2 n = 4x = 36) ‘Key’ lime + ‘Valencia’ orange, and between diploid ‘Wilking’ and ‘Fortune’ mandarins and an autotetraploid ‘Dancy’ mandarin (2 n = 4x = 36). To understand the opportunity to employ them in further backcross programs, the cytological mechanisms underlying their ploidy level were unambiguously determined using six SSR primers. PCR conditions were optimized and skewness in template/product ratios were verified. Tetraploid allelic configurations were determined from PCR based dosage effects using electropherogram peak heights to estimate the copy number per allele. In all the tetraploid hybrids we found out that diploginy (2 n eggs) has occurred, contributing the extra haploid genome in the tetraploids. According to the marker genotypes, it was further inferred that the 2 n eggs in ‘Femminello’ lemon resulted from first division restitution (FDR), while in ‘Wilking’ and ‘Fortune’ mandarins 2 n eggs occurred in second division restitution (SDR). These new genotypes, with their improved genetic female background, can be therefore considered very valuable in our citrus genetic improvement program as pollen donors in backcrosses suitable to eliminate negative traits. [ABSTRACT FROM AUTHOR]

'Milam' (a purported hybrid of Citrus jambhiri Lush) + 'Femminello' lemon (Citrus limon L. Burm. f.) allotetraploid somatic hybrids were used as pollen parents in interploid crosses with diploid 'Femminello' lemon to achieve mal secco tolerance in different populations of seedless triploid lemon types with good fruit quality. A total of 137 plantlets were obtained and subjected to screening experiments, in order to distinguish zygotic embryos from nucellars. Here we report on and discuss the results obtained with three techniques: flow cytometry, isozyme analysis and ISSR-PCR (the inter-simple sequence repeats-polymerase chain reaction). ISSR-PCR resulted to be a very efficient and reliable technique for the identification of zygotic plantlets.