Amplified DNA Fingerprinting Research Articles

Field performance evaluation and genetic integrity assessment of cryopreserved papaya clones

This paper is the first report of field performance and evaluation of morphological traits following cryopreservation in four genotypes of Carica papaya (Z6, 97, TS2 and 35). It also describes the successful establishment of in vitro plantlets following vitrification-based cryopreservation of shoot tips and their acclimatisation through to field establishment. Cloned plants resulting from untreated controls, as well as controls taken at three other stages of the cryopreservation process (dissection, pre-treatment, plant vitrification solution 2 (PVS2) treatment) and cryopreserved plants were established to ensure a rigorous appraisal of any variation. Results indicate no differences between any of the control plants or cryopreserved plants for either growth performance or morphology. In addition, both randomly amplified DNA fingerprinting and amplified DNA methylation polymorphism markers were used to assess any genomic or methylation changes in genotype 97 at four different developmental stages post cryopreservation (in vitro, acclimatisation and field). Only small genomic DNA modifications (0-8.3%) were detected in field stage plants and methylation modifications (0-4.3%) were detected at both the in vitro and field stages for samples treated with PVS2 or cryopreservation.

Establishment and characterization of a new Aedes aegypti (L.) (Diptera: Culicidae) cell line with special emphasis on virus susceptibility

A new cell line from the neonate larvae of Aedes aegypti (L) mosquito was established and characterized. The cell line at the 50th passage (P) level consisted of three prominent cell types, i.e., epithelial-like cells (92%), fibroblast-like cells (7%), and giant cells ( approximately 1%). Karyological analysis showed diploid (2n = 6) number of chromosomes in >75% cells at P-50. The growth kinetics studied at 52nd passage level showed approximately tenfold increase in cell number over a 10-d study period. The species specificity studies using DNA amplification fingerprinting profile analysis using RAPD primers demonstrated 100% homology with the host profile showing the integrity of the cell line. Electron microscopy revealed the absence of mycoplasma or other adventitious agents. The cell line supported the multiplication of seven arboviruses, i.e., Chikungunya (CHIK), Japanese encephalitis, West Nile, dengue 2 (DEN-2), Chandipura, vesicular stomatitis, and Chittoor viruses. The cell line did not replicate Ganjam and Kaisodi viruses. CHIK virus yield in the new cell line was approximately 3log and 0.5log 50% tissue culture infective dose (TCID(50))/mL higher than Vero E6 and C6/36 cell lines, respectively. In the case of DEN-2 virus, it yielded 1log TCID(50)/mL higher than Vero E6, but lesser than C6/36 cell line. Due to its high susceptibility to a broad spectrum of viruses, the new cell line may find application in virus isolation during epidemics and in antigen production.

Development of taxon‐specific sequence characterized amplified region (SCAR) markers based on actin sequences and DNA amplification fingerprinting (DAF): a case study in the Phoma exigua species complex

Phoma exigua is considered to be an assemblage of at least nine varieties that are mainly distinguished on the basis of host specificity and pathogenicity. However, these varieties are also reported to be weak pathogens and secondary invaders on non-host tissue. In practice, it is difficult to distinguish P. exigua from its close relatives and to correctly identify isolates up to the variety level, because of their low genetic variation and high morphological similarity. Because of quarantine issues and phytosanitary measures, a robust DNA-based tool is required for accurate and rapid identification of the separate taxa in this species complex. The present study therefore aims to develop such a tool based on unique nucleotide sequence identifiers. More than 60 strains of P. exigua and related species were compared in terms of partial actin gene sequences, or analysed using DNA amplification fingerprinting (DAF) with short, arbitrary, mini-hairpin primers. Fragments in the fingerprint unique to a single taxon were identified, purified and sequenced. Alignment of the sequence data and subsequent primer trials led to the identification of taxon-specific sequence characterized amplified regions (SCARs), and to a set of specific oligonucleotide combinations that can be used to identify these organisms in plant quarantine inspections.

A cell strain cloned from Spodoptera exigua cell line (IOZCAS-Spex-II) highly susceptible to S. exigua nucleopolyhedrovirus infection

A cell strain (IOZCAS-Spex-II-A) cloned from IOZCAS-Spex-II, a cell line established from the fat body of Spodoptera exigua (Lepidoptera: Noctuidae) larva, was characterized, and its capability to produce S. exigua nucleopolyhedrovirus was high with infection rate exceeding 90% compared with its parental cell line IOZCAS-Spex-II that scored only 50%. Growth curve of budded virus (BV) in the strain was analyzed and the titer of BV reached the highest of 3.7 x 10(4) pfu/mL by 96 h after inoculation. Concentration of occlusion bodies (OBs) produced by the cloned cell strain (IOZCAS-Spex-II-A) was 7.1 x 10(7) OBs/mL, while the parental cell line produced 2.4 x 10(7) OBs/mL. The average yield of the virus was 176 OBs/cell of IOZCAS-Spex-II-A compared with 211 OBs/cell that of the parental cell line. Significant differences were observed in virus production, growth characters, cell shape, between the parental cell line, and its clone. The cell lines (IOZCAS-Spex-II and IOZCAS-Spex-II-A) were also susceptible to Autographa californica multiple nucleopolyhedrovirus infection. In addition, they were characterized with regard to their growth rates and DNA amplification fingerprinting technique employing polymerase chain reaction.

DNA amplification fingerprinting analysis of genetic variation withinFusarium oxysporumf.sp.zingiberi

Genetic variation among 29 isolates of Fusarium oxysporum f.sp. zingiberi (Foz) collected from diseased ginger rhizome in production regions throughout Queensland was analysed using DNA amplification fingerprinting (DAF). Eight isolates of other Fusarium species and/or formae speciales were included for comparative analysis. Within the Foz isolates, three haplotypes were identified based on 17 polymorphic bands generated with five primers. Two groups showed very little genetic variation (98.6% similarity), whereas the third single isolate was quite distinct in terms of its molecular profile (77.2% similarity). Genetic similarity among the Fusarium solani, F. oxysporum f.sp. lycopersici and F. oxysporum f.sp. cubense races 1, 3 and 4 isolates compared well with the published literature.

Molecular Identification Keys for Cultivars and Lines of Cornus florida and C. kousa Based on Simple Sequence Repeat Loci

Flowering dogwood (Cornus florida) and kousa dogwood (C. kousa) are popular ornamental species commonly used in the horticultural industry. Both trees are valued for their beautiful floral display and four-season appeal. Species-specific simple sequence repeat (SSR) loci were used to genotype and assess genetic diversity of 24 flowering dogwood cultivars and breeding lines and 22 kousa dogwood cultivars. Genetic diversity was determined by allele sharing distances and principal coordinate analysis and was high in both species. Molecular identification keys were developed for cultivars and breeding lines of each species using a few polymorphic SSRs loci (four in C. florida and five in C. kousa). Most (18 of 24) of the flowering dogwood and all (22 of 22) kousa dogwood accessions could be distinguished from each other using these SSRs; those that could not were resolved using DNA amplification fingerprinting. The reliability of both keys was assessed using five anonymous cultivars for each dogwood species, which were correctly identified using the molecular keys. The genetic information presented here will be useful for identification and verification of cultivars for nurseries and as molecular markers for breeders and researchers.

Low Genetic Diversity in the Ground Parrot (Pezoporus wallicus) Revealed by Randomly Amplified DNA Fingerprinting

The ground parrot (Pezoporus wallicus) is a vulnerable species that occurs in isolated pockets of heathland and sedgeland of Australia. This study used randomly amplified DNA fingerprinting (RAF) to examine genetic diversity in the eastern population of the ground parrot. The seven primers used produced an average of 68 markers per primer, and the number of unambiguous polymorphic markers per primer averaged 6.3 (9.2%). Overall genetic similarity was 0.978 ± 0.03. The low level of genetic diversity revealed by RAF is comparable to the lower end of diversity found in species that are declared endangered.

Assessment of genetic and epigenetic changes following cryopreservation in papaya

A vitrification based cryopreservation technique for storage of in vitro shoot tips of papaya has been tested to ensure applicability across a range of genotypes and to assess the stability of both genotype and phenotype of such clonal material following cryopreservation. Shoot tips of 12 genotypes were cryopreserved, recovery rates were determined and resultant plants were screened for genetic and epigenetic changes. Genomic DNA structure was explored using polymerase chain reaction (PCR) based randomly amplified DNA fingerprinting (RAF), and methylation patterns were monitored using the amplified DNA methylation polymorphism (AMP) PCR technique. Plantlets were recovered following cryopreservation in all but one genotype and recovery rates of 61-73% were obtained from six genotypes. The regenerated plantlets showed varying levels of genomic DNA modifications (0-10.07%), and methylation modifications (0.52-6.62%) of detected markers. These findings have not been reported previously for papaya, and indicate some genotype dependent variability in DNA modifications occur following cryopreservation which may result in somaclonal variation.

IDENTIFICATION OF CERTAIN PAPAYA CULTIVARS AND SEX IDENTIFICATION IN PAPAYA BY DNA AMPLIFICATION FINGERPRINTING (DAF)

DNA amplification fingerprinting (DAF) was employed to study the relationships among fourteen cultivars of the papaya (Carica papaya L.) collected from Srisaket Horticultural Research Center and Chantaburi Horticultural Research Center. Ten-base synthetic oligonucleotides were chosen that gave multiple PCR amplification products using papaya DNA as template. Using the 11 primers, a total of 129 distinct fragments were amplified. Cultivars were scored for the presence or absence of DAF fragments and grouped by cluster analysis using sample matching coefficients of similarity and the unweighted pair-group method with arithmetic average (UPGMA). The results identified three groups of cultivars. The first group was comprised of 'Pak Chong', 'Khaeg Dum', 'Mexico Mammy' and 'Khaeg Nuan'. The second group was comprised of 'Cocoa KarnDum', 'Apple', 'Tainung', 'Mae Hia', 'Florida tolerant', 'Sunset' and 'Mexico Amerilla'. The third group was comprised of 'Taiwan', 'Kapoho Solo' and 'Richter'. The DAF technology is a rapid, precise and sensitive technique for genomic analysis, and is recommended as an accurate method to identify plant genotypes within the same species. The best DAF reaction mixture contained 30 ng of genomic DNA, 5 mM magnesium ion, 2.5 mM primer per 20 μl reaction. A total of 52 primers were used in bulk segregate analysis (BSA) against of male, female and hermaphroditic plants. The OPA 06 (5' GGT CCC TGAC 3') primer could be used to identify the sex type of papaya plants. This primer produced two polymorphic bands: one of ∼ 365 base pairs (bp) from hermaphrodite bulk DNA and the other of ∼ 360 bp from the male bulk DNA. Neither band was detected for females. Evaluation of the accuracy of OPA 06 analysis was done using 254 plants of different generations and their original parents, and the analysis correctly identified sex type for 88.18% of the plants. In the final experiment, 47 tissue cultured hermaphrodite plants of 'Khaeg Dum' were examined using OPA 06, and the sex type was identified correctly for 100% of the plants.

Identification of Insect Cell Lines from 8 Lepidopteran species by DNA Amplification Fingerprinting

Identification of Insect Cell Lines from 8 Lepidopteran species by DNA Amplification Fingerprinting

Genetic diversity and phylogenetic relationships in Vigna Savi germplasm revealed by DNA amplification fingerprinting

The pantropical genus Vigna (Leguminosae) comprises 7 cultivated species that are adapted to a wide range of extreme agroclimatic conditions. Few data are available on the relationships among these cultivated species or on their importance as sources of resistance against biotic and abiotic stresses. Therefore, we optimized DNA amplification fingerprinting (DAF) to estimate the genetic diversity within, and genetic relationships among, a representative core collection of cowpea, as compared with 16 accessions representing cultivars from 6 Vigna species. A set of 26 primers was selected from 262 tested random primers and used for the characterization of 85 Vigna accessions (6 V. angularis, 4 each of V. mungo and V. radiata, 2 V. umbellata, 1 V. aconitifolia, and 68 V. unguiculata), with Phaseolus vulgaris subsp. vulgaris as outgroup. A total of 212 polymorphic bands were used for maximum parsimony analysis. Our results clearly distinguished Brazilian from African V. unguiculata genotypes. At the species level, V. angularis was the most related and V. radiata the most divergent species relative to V. unguiculata. DAF markers were also informative at the intraspecific level, detecting a large diversity between cowpea cultivars. The implications of the presented results for cowpea breeding programs are discussed.

IDENTIFICATION OF CERTAIN PAPAYA CULTIVARS AND SEX IDENTIFICATION IN PAPAYA BY DNA AMPLIFICATION FINGERPRINTING (DAF)

DNA amplification fingerprinting (DAF) was employed to study the relationships among fourteen cultivars of the papaya (Carica papaya L.) collected from Srisaket Horticultural Research Center and Chantaburi Horticultural Research Center. Ten-base synthetic oligonucleotides were chosen that gave multiple PCR amplification products using papaya DNA as template. Using the 11 primers, a total of 129 distinct fragments were amplified. Cultivars were scored for the presence or absence of DAF fragments and grouped by cluster analysis using sample matching coefficients of similarity and the unweighted pair-group method with arithmetic average (UPGMA). The results identified three groups of cultivars. The first group was comprised of 'Pak Chong', 'Khaeg Dum', 'Mexico Mammy' and 'Khaeg Nuan'. The second group was comprised of 'Cocoa KarnDum', 'Apple', 'Tainung', 'Mae Hia', 'Florida tolerant', 'Sunset' and 'Mexico Amerilla'. The third group was comprised of 'Taiwan', 'Kapoho Solo' and 'Richter'. The DAF technology is a rapid, precise and sensitive technique for genomic analysis, and is recommended as an accurate method to identify plant genotypes within the same species. The best DAF reaction mixture contained 30 ng of genomic DNA, 5 mM magnesium ion, 2.5 mM primer per 20 μl reaction. A total of 52 primers were used in bulk segregate analysis (BSA) against of male, female and hermaphroditic plants. The OPA 06 (5' GGT CCC TGAC 3') primer could be used to identify the sex type of papaya plants. This primer produced two polymorphic bands: one of ∼ 365 base pairs (bp) from hermaphrodite bulk DNA and the other of ∼ 360 bp from the male bulk DNA. Neither band was detected for females. Evaluation of the accuracy of OPA 06 analysis was done using 254 plants of different generations and their original parents, and the analysis correctly identified sex type for 88.18% of the plants. In the final experiment, 47 tissue cultured hermaphrodite plants of 'Khaeg Dum' were examined using OPA 06, and the sex type was identified correctly for 100% of the plants.

INTERGENERIC HYBRIDISATION BETWEEN CARICA PAPAYA AND WILD VASCONCELLEA SPECIES AND IDENTIFICATION OF A PRSV-P RESISTANCE GENE

Intergeneric hybridisations between Carica papaya and related Vasconcellea species have been conducted for 15 years, despite limitations due to postzygotic barriers. Successful crosses have been achieved between C. papaya and PRSV-P resistant Vasconcellea species cauliflora and pubescens, however the infertility of these hybrids has prevented further backcrossing. A few hybrids with low fertility have been present in large populations of C. papaya x V. quercifolia. An extensive backcrossing program between these hybrids and papaya is currently underway in both Australia and the Philippines. A single dominant gene for PRSV-P resistance in V. pubescens has been mapped by use of dominant, polymorphic randomly amplified DNA fingerprint (RAF) markers in an F2 interspecific population of V. parviflora (PRSV-P susceptible) and V. pubescens (PRSV-P resistant). Using a bulked segregant analysis strategy, a RAF marker was identified as being linked to the resistant phenotype and was shown to be present in other PRSV-P resistant Vasconcellea species. It mapped to within 6.3 cM of he predicted PRSV-P resistance locus. The RAF marker was converted into a co-dominant CAPS marker, diagnostic for resistance based on digestion with the restriction endonuclease Psi I. This marker may provide a useful tool for future marker-assisted breeding programs aimed at introgressing PRSV-P resistance genes from wild Vasconcellea species into the commercially important papaya.

The extent of clonality in large stands of Lycopodium annotinum L.

In the Rothaargebirge (Northrhine-Westphalia) the interrupted club moss ( Lycopodium annotinum L.) often grows in large (maximum size of patches observed: 18,000 m 2) and dense (club moss cover 60–90%) clusters. By DNA amplification fingerprinting (DAF) analysis it was shown in three populations comprising a total of 69 samples that the clusters are of great genetical uniformity. Based on data from 47 polymorphic loci amplified with 13 primers 11 multilocus genotypes were identified, nine in one population and one per population in the other two. The data were subjected to similarity analysis, generating Jaccard′s similarity coefficients, on the basis of which cluster analysis was performed. Calculation of Nei's genetic diversity (1987) resulted in a mean value of 0.77 over all samples and a mean within populations diversity of 0.3. Corrected G′ ST value for the fraction of diversity among the populations was 0.7. Two out of three sites are most probably formed by one clone only. As the annual growth rate of the shoots is known, it has to be stated that the stands under investigation are at least 80–90 years old. Taking into consideration that annual accretion is virtually non-existent following forest thinning measures, it becomes very likely that the club moss plants are approximately as old as the trees of their forests (150–180 years). This hypothesis is supported by the observation that within the forests only large (i.e. old) stands, but no single shoots (i.e. young individuals) of the club moss exist. Thus one can conclude that in undisturbed forests no sexual reproduction of L. annotinum takes place.

Diosgenin Contents and DNA Fingerprint Screening of Various Yam (Dioscorea sp.) Genotypes

In addition to the importance of many Dioscorea species (yams) as starchy staple food, some representatives are known and still used as a source for the steroidal sapogenin diosgenin, which, besides phytosterols derived from tall-oil, is an important precursor for partial synthesis of steroids for pharmaceutical research and applications. While in edible yams the diosgenin content should be as low as possible, a high yield of the compound is preferable for cultivars which are grown for the extraction of sterols. In the past, miscalculations and insufficiently precise techniques for quantification of diosgenin prevailed. Therefore we set out to re-evaluate the steroid content of a world collection of Dioscorea species, using leaves as sample material. We optimized diosgenin quantification techniques and fingerprinted the whole collection with the DNA amplification fingerprinting (DAF) technique. Total diosgenin contents ranged from 0.04 to 0.93% of dry weight within the collection. Several Dioscorea cultivars can be characterized via their DAF fingerprint patterns.

APPLICATION OF SCAR MARKERS LINKED TO A PUTATIVE PRSV-P RESISTANCE LOCUS FOR ASSISTED BREEDING OF RESISTANT C. PAPAYA CULTIVARS

Papaya ringspot virus type P (PRSV-P) is a serious disease of Carica papaya. Several wild relatives of the papaya, belonging to the genus Vasconcellea, exhibit complete resistance to PRSV-P, namely V. cundinamarcensis, V. cauliflora and V. quercifolia. Molecular genetic analysis of interspecific hybrids of V. cundinamarcensis and V. parviflora using Randomly Amplified DNA Fingerprinting (RAF) permitted the construction of genetic maps of these species. Identification and mapping of several DNA markers linked to a PRSV-P resistance gene locus in V. cundinamarcensis was possible, two of which (OPK41R and OPA115R), resulted from a Single Nucleotide Polymorphism (SNP), in close proximity to the PRSV-P resistance gene locus. The Sequence Characterised Amplified Regions (SCAR's), were shown to be present in both parents, and produced codominant markers, which were applied as reliable diagnostic markers for resistant and susceptible genotypes. The OPK41R SCAR was identified in 4 additional Vasconcellea species made available, including V. cauliflora and V. quercifolia, but not in C. papaya. The indel identified may be useful for the selection of PRSV-P resistant individuals in breeding programs with Vasconcellea and C. papaya to assist introgression of putative resistance genes. The markers were subsequently applied to an intergeneric hybrid population from C. papaya and V. quercifolia. Genetic characterisation of the hybrid population using RAF markers confirmed hybridity of the F1 and reduced V. quercifolia DNA content of back cross generations. The OPK41R and OPA115R SCARs were tested in the quercifolia hybrids, and did not identify linkage to PRSV-P resistance. From the results it can be inferred that Vasconcellea may contain multiple sources of resistance to PRSV-P. Overall these studies indicate that there is significant potential for the continued application of DNA markers in molecular breeding for PRSV-P resistant papaya genotypes.

Population Biology of Pseudoperonospora humuli in Oregon and Washington.

Pseudoperonospora humuli populations from Oregon and Washington were analyzed for genetic variation using random amplified polymorphic DNA (RAPD) and DNA amplification fingerprinting (DAF) markers. The genetic structure of the Oregon and Washington populations differed considerably. There was little genetic diversity in Washington, with only five RAPD and six DAF groups detected among 40 isolates tested. One genotype was predominant in Washing-ton. In contrast, 18 RAPD and 34 DAF groups were found among the 40 isolates tested from Oregon. No unique band profile associated with host cultivar was observed. It is suggested that the distinct difference in population structure between the two geographic regions might be due to climatic differences resulting in a higher frequency of sexual reproduction of P. humuli in Oregon than in Washington.

Development of a codominant CAPS marker linked to PRSV-P resistance in highland papaya

Development of resistant papaya varieties is widely considered the best strategy for long-term control of the papaya ringspot virus type P (PRSV-P). Several species of "highland papaya" from the related genus Vasconcellea exhibit complete resistance to PRSV-P, and present a valuable source of resistance genes with potential for application in Carica papaya. The objectives of this study were two fold; to identify molecular markers linked to a previously characterised PRSV-P resistance gene in V. cundinamarcensis (psrv-1), and to develop codominant marker based strategies for reliable selection of PRSV-P resistant genotypes. Using a bulked segregant analysis approach, dominant randomly amplified DNA fingerprint (RAF) markers linked to prsv-1 were revealed in the resistant DNA bulk, which comprised F2 progeny from a V. parviflora (susceptible) x V. cundinamarcensis (resistant) interspecific cross. One marker, Opk4_1r, mapped adjacent to the prsv-1 locus at 5.4 cM, while a second, Opa11_5r, collocated with it. Sequence characterisation of the Opk4_1r marker permitted its conversion into a codominant CAPS marker (PsiIk4), diagnostic for the resistant genotype based on digestion with the restriction endonuclease PsiI. This marker mapped within 2 cM of the prsv-1 locus. Psilk4 was shown to correctly identify resistant genotypes 99% of the time when applied to interspecific F2 progeny segregating for the resistant character, and has potential for application in breeding programs aimed to deliver the PRSV-P resistance gene from V. cundinamarcensis into C. papaya.

Comparison of Aflatoxigenic and Nonaflatoxigenic Isolates of Aspergillus flavus using DNA Amplification Fingerprinting Techniques

Aspergillus flavus is a filamentous fungus that produces mycotoxins in many food and feed crops, such as maize (Zea mays L.). Isolates were analyzed for toxin production by nucleic acid profiles in an attempt to differentiate aflatoxigenic from nonaflatoxigenic isolates. A total of 41 aflatoxigenic and 34 nonalfatoxigenic isolates were included in the study. The isolates were evaluated initially using DNA amplification fingerprinting (DAF) without clear resolution of the groups. A weak association of aflatoxigenic isolates was observed, as evidenced by their clustering in 18 of 81 trees recovered from maximum parsimony analysis of binary characters derived from arbitrary signatures from amplification profiles (ASAP) data; nonaflatoxigenic isolates exhibited a pattern of paraphyletic laddering. Up to five markers unambiguously supported the aflatoxigenic isolate grouping, but the presence of alternative conflicting topologies in equally parsimonious trees precluded the observation of meaningful statistical support. With additional markers for genome of A. flavus, this method could be used to resolve toxigenic from nontoxigenic strains. This additional work could resolve aflatoxigenic isolates of A. flavus present on maize plants using ASAP, which would reduce labor intense costs and potentially lead to faster determination of resistant cultivars in breeding efforts.

Evidence of a Genomic Biomarker in Normal Human Epithelial Mammary Cell Line, MCF‐10A, That Is Absent in the Human Breast Cancer Cell Line, MCF‐7

This study investigated the use of DNA amplification fingerprinting (DAF) to identify biomarkers useful in the elucidating genetic factors that lead to carcinogenesis. The DNA amplification fingerprinting (DAF) technique was used to generate fingerprint profiles of a normal human mammary epithelial cell line (MCF-10A) and a human breast cancer cell line (MCF-7). When compared with one another, a polymorphic biomarker gene (262 base pairs (bps)) was identified in MCF-10A but was not present in MCF-7. This gene was cloned from the genomic DNA of the MCF-10A cell line, and subjected to Genbank database analysis. The analysis of the nucleotide sequence polymorphic marker (Genbank account: AC079630) shows that this biomarker has 100% homology with the nucleotide sequence of human chromosome 12 BAC RP11-476D10 (bps 19612-19353). The nucleotide sequence was used for possible protein translation product and the result obtained indicated that the gene codes for hypothetical protein XF2620. In order to evaluate the effects that the 262 bps biomarker would have on the morphology of MCF-7 cells, it was transfected into MCF-7 cells. There were observable changes in the morphology of the transfected cells. These changes included an increase in cell elongation and a decrease in cell aggregation.