Elevated CISD2 expression predicts poor diagnosis and promotes invasion and migration of prostate cancer cells.

OBJECTIVE: To explore roles of CDGSH iron-sulfur domain-containing protein 2 (CISD2) in the progression of prostate cancer (PCa) cells, and relationships between CISD2 expression and the prognosis and clinical pathological parameters in PCa patients. PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) analysis and Western blot analysis were used to detect the CISD2 expression in PCa tissues and cells. CISD2 siRNA was used to inhibit the CISD2 expression. Kaplan-Meier method and Log rank analysis were performed to determine survival analysis while Chi-square test was performed to analyze the association between CISD2 and clinicopathological parameters of PCa patients. Transwell assay and wound healing assay was conducted to examine the invasion and migration ability of PCa cells, respectively. RESULTS: CISD2 was up-regulated in PCa tissues and cells, and showed positive association with the poor prognosis, T stage, lymphatic invasion, prostate-specific antigen (PSA) level, and distant metastasis of PCa patients. Besides, we found that inhibition of CISD2 significantly impaired the migration and invasion ability of PCa cells. CONCLUSIONS: The paper demonstrated that CISD2 could act as a new target for the diagnosis and treatment of PCa patients.

Interferon-γ affects leukemia cell apoptosis through regulating Fas/FasL signaling pathway.

OBJECTIVE: Imbalance of hematopoietic cell proliferation and apoptosis is one of the major causes of leukemia. Enhanced cell proliferation and reduced apoptosis lead to hemocytes accumulation. Fas/FasL signaling pathway promotes cell apoptosis. This study investigated the impact of interferon γ (IFN-γ) on chronic myelogenous leukemia cell proliferation and apoptosis to elucidate its interaction with Fas/FasL signaling pathway. PATIENTS AND METHODS: Leukemia K562 cells were routinely cultivated and treated with 10 U/ml, 100 U/ml, and 1000 U/ml interferon for 12 h, 24 h, and 48 h, respectively. MTT assay was applied to test cell proliferation. TUNEL assay was adopted to determine cell apoptosis. Western blot was selected to detect Fas/FasL expression. RESULTS: Different concentrations of IFN-γ inhibited cell proliferation at various time points. IFN-γ at 1000 U/ml treatment for 48 h exhibited the strongest suppressive effect on cell proliferation (p < 0.05). IFN-γ intervention enhanced K562 cell apoptosis with concentration and time dependence (p < 0.05). Fas and FasL proteins expressions upregulated after treated by IFN-γ following dose elevation and time extension (p < 0.05). CONCLUSIONS: IFN-γ inhibits leukemia K562 cell proliferation and promotes cell apoptosis via facilitating Fas and FasL proteins expressions.

FERMT1 mediates epithelial-mesenchymal transition to promote colon cancer metastasis via modulation of ß-catenin transcriptional activity.

We previously demonstrated that fermitin family member 1 (FERMT1) was significantly overexpressed in colon cancer (CC) and associated with poor metastasis-free survival. This study aimed to investigate the precise role of FERMT1 in CC metastasis and the mechanism by which FERMT1 is involved in the epithelial-mesenchymal transition (EMT). Correlations between FERMT1 and EMT markers (E-cadherin, Slug, N-cadherin and ß-catenin) were examined via immunohistochemistry in a cohort of CC tissues and adjacent normal colon mucosae. A series of in vitro and in vivo assays were performed to elucidate the function of FERMT1 in CC metastasis and underlying mechanisms. The upregulated expression of FERMT1 in CC tissues correlated positively with that of Slug, N-cadherin and ß-catenin, but correlated inversely with E-cadherin expression. Altered FERMT1 expression led to marked changes in the proliferation, migration, invasion and EMT markers of CC cells both in vitro and in vivo. Investigations of underlying mechanisms found that FERMT1 interacted directly with ß-catenin and activated the Wnt/ß-catenin signaling pathway by decreasing the phosphorylation level of ß-catenin, enhancing ß-catenin nuclear translocation and increasing the transcriptional activity of ß-catenin/TCF/LEF. Activation of the Wnt/ß-catenin pathway by CHIR99021 reversed the effect of FERMT1 knockdown, whereas inhibition of the Wnt/ß-catenin pathway by XAV939 impaired the effect of FERMT1 overexpression on EMT and cell motility. In conclusion, findings of this study suggest that FERMT1 activates the ß-catenin transcriptional activity to promote EMT in CC metastasis.

Molecular cloning and characterization of lymphocyte cell kinase from humphead snapper (Lutjanus sanguineus).

Lymphocyte cell kinase (LCK) belongs to the Src family of tyrosine kinases, which involves in the proliferation control of lymphocytes. In this study, we cloned the LCK gene of humphead snapper (Lutjanus sanguineus) (designed as LsLCK). Sequence analysis showed that the full-length cDNA of LsLCK was 2279 bp, contained a 1506-bp open reading frame (ORF), encoding a polypeptide of 501 amino acids. The deduced amino acid possessed the typical structural features of known LCK proteins, including four Src homology (SH) domains arranged as the SH1 domain followed by a regulatory C-terminal tail (COOH-domain), SH2 and SH3 adapter domains and SH4 domain which required for membrane attachment and CD4/CD8 binding. Fluorescent quantitative real-time PCR analysis indicated that LsLCK transcripts were expressed mainly in thymus, spleen and head kidney in healthy fish. Moreover, the mRNA expressions in these tissues were significantly up-regulated after challenge with Vibrio harveyi. The results of immunohistochemistry showed that LsLCK protein localized distinctly in cytoplasm of cell in thymus, spleen and head kidney. Taken together, these findings indicated that LsLCK may play an important role in the immune response of humphead snapper against bacterial infection.

MiRNA-621 sensitizes breast cancer to chemotherapy by suppressing FBXO11 and enhancing p53 activity.

MicroRNAs (miRNAs) have been demonstrated to have critical roles in regulating cancer cell proliferation, survival and sensitivity to chemotherapy. The potential application of using miRNAs to predict therapeutic response to cancer treatment holds high promise, but miRNAs with predictive value remain to be identified and underlying mechanisms have not been completely understood. Here, we show a strong correlation between miR-621 expression and chemosensitivity to paclitaxel plus carboplatin (PTX/CBP) regimen, an effective neoadjuvant chemotherapy for breast cancer patients. High level of miR-621 predicts better response to PTX/CBP regimen neoadjuvant chemotherapy in breast cancer patients, who also tend to achieve pathological complete response. Ectopic overexpression of miR-621 promoted apoptosis and increased chemosensitivity to PTX and CBP both in cultured breast cancer cells and in xenograft tumor model. We further show that FBXO11 is a direct functional target of miR-621 and miR-621 level is negatively correlated with FBXO11 expression in breast cancer patients. Ectopic expression of FBXO11 attenuated increased apoptosis in breast cancer cells overexpressing miR-621 upon PTX or CBP treatment. Consistently, high FBXO11 expression significantly correlated with poor survival in breast cancer patients. Mechanistically, we found in breast cancer cells FBXO11 interacts with p53 and promotes its neddylation, which suppressed the p53 transactivity. Accordingly, miR-621-dependent FBXO11 suppression enhanced p53 activity and increased apoptosis in breast cancer cells exposed to chemotherapeutics. Taken together, our data suggest that miR-621 enhances chemosensitivity of breast cancer cells to PTX/CBP chemotherapy by suppressing FBXO11-dependent inhibition of p53. miR-621 may serve as a predictive biomarker and a potential therapeutic target in breast cancer treatment.

Variation of craniocervical junction volume as an effective parameter for basilar invagination treatment.

OBJECTIVE: The major pathological change in basilar invagination (BI) is represented in the decrease of craniocervical junction (CVJ) volume resulting from abnormal bone protrusion around the foramen magnum. The diagnosis and clinical evaluation of BI is mainly based on the clinical manifestations and radiographic measurements by means of calculation of the scan lines of CVJ in X-ray, CT and MRI. With the transoral decompression atlantoaxial reduction plate (TARP III) system, the decompression, reduction and fixation can be achieved to decompress and stabilize medulla spinalis change the position of the dens in CVJ, thus expand the CVJ relative volume, relieve the compression on medulla spinalis and the nerve injury. However, the correlation between the dens position change and the variation of CVJ has not been established previously. This study focused on the clinical significance of the variation of craniocervical junction (CVJ) volume caused by the dens position change for the treatment of BI. PATIENTS AND METHODS: We've performed an analysis of data from 62 BI patients admitted from January 2008 to May 2013, who were treated by TARP III system. The data include preoperative, postoperative JOA scores (Japanese Orthopaedic Association scores, 17 points method), preoperative and postoperative X-ray, thin-slice CT scan with three-dimensional reconstruction and MRI scan to measure the cervicomedullary angle (CMA). We have analyzed the preoperative and postoperative three-dimensional CT data by means of MIMICS 10.01 software system according to the Box volume (BV) method to determine the changes of CVJ volume resulting from preoperative and postoperative dens position change, assessed the correlation between the CVJ volume changes and the JOA scores with correlation between CMA change and the JOA scores. All data were analyzed by paired t-test and Pearson correlation analysis. RESULTS: In all 62 patients, JOA scores were recovered from preoperative 9.26 ± 1.66 to postoperative 13.02 ± 1.44, CMA change rate was 21%, and CVJ volume change rate was 36%. The CMA change rate and the JOA score recovery rat exhibited relevance, as Pearson's correlation coefficient was 0.46 (p < 0.005). The Pearson's correlation coefficient between CVJ volume change rate and JOA score recovery rate was 0.63 (p < 0.005), and the CVJ volume change rate was significantly different while compared with the correlation between CMA change rate and JOA score (p < 0.005). CONCLUSIONS: the CVJ volume change rate is a sensitive and reliable parameter for the evaluation of neurological function improvement in patients with BI. It can be used as a predictor to evaluate the postoperative neurological recovery.

Analysis of carcass and meat quality traits and nutritional values of hybrid wild boars under different crossing systems.

The aim of this study was to analyze the results of two crossing systems between wild boars and different domesticated pig breeds. Hybrid wild boars were produced by crossing captured wild boars with Meishan pigs and LY sows according to the traditional production system. The resultant commercial hybrids were black and white in coat color, respectively. Significant differences were found in the carcass and meat quality traits and nutritional values between these two hybrid wild boars. Compared with the white hybrid wild boars, at the age of 300 days, the body weight of black hybrid wild boars was 9.41 kg lower, while percent lean was 2.51% less and percent fat 2.45% higher (P < 0.05). The black hybrid wild boars had higher pH2 (6.17 vs 6.09) and intramuscular fat (3.34 vs 2.52%), lower drip loss (2.21 vs 2.68%) and shear force (44.00 vs 52.23) (P < 0.05), and more unsaturated fatty acids and essential amino acids (P < 0.05). In conclusion, cross breeding was shown to be an effective method to improve the overall production performance of wild boars, but crossing with different dam line breeds caused different responses. Compared with the white hybrid wild boars, the black hybrid wild boars had worse growth rate and carcass traits, but better meat quality traits and nutritional values.

Membrane microparticles and diseases.

Membrane microparticles (MPs) are plasma membrane-derived vesicles shed by various types of activated or apoptotic cells including platelets, monocytes, endothelial cells, red blood cells, and granulocytes. MPs are being increasingly recognized as important regulators of cell-to-cell interactions. Recent evidences suggest they may play important functions not only in homeostasis but also in the pathogenesis of a number of diseases such as vascular diseases, cancer, infectious diseases and diabetes mellitus. Accordingly, inhibiting the production of MPs may serve as a novel therapeutic strategy for these diseases. Here we review recent advances on the mechanism underlying the generation of MPs and the role of MPs in vascular diseases, cancer, diabetes, inflammation, and pathogen infection.

Effect of muscle-fiber type on glycogenin-1 gene expression and its relationship with the glycolytic potential and pH of pork.

This study analyzed the effect of muscle-fiber type composition on glycogenin-1 (GYG) gene expression and its impact on pH. The longissimus dorsi (LD) muscle contains more type IIB fibers (75.10%) than does the psoas major (PM) muscle (41.58%), while the PM has more type I (3.65 vs 0.94%), type IIA (34.15 vs 10.63%), and type IIX (20.62 vs 13.33%) fibers. Compared with PM, glycolytic potential (GP), pH45 min, and ΔpH from 45 min to 24 h post-mortem were all relatively higher in LD. Glycogen metabolites (lactate and GP) were negatively correlated with pH24 h and positively correlated with ΔpH. Expression of GYG was generally higher in LD. GYG expression was positively correlated with glycogen metabolite (lactate and GP) content and ΔpH, and was negatively correlated with pH24 h. These data confirm that the muscle-fiber type and GP have significant effects on ultimate pH and pH decline, and suggest that expression of GYG in muscles is related to the metabolism of glycogen and may impact GP, ΔpH, and ultimate pH. High expression of GYG was associated with a high glycogen content, large pH decline, and low ultimate pH in muscles post-mortem.

Effects of different activation regimens on pronuclear formation and developmental competence of in vitro-matured porcine oocytes after intracytoplasmic sperm injection.

To improve the activation protocol for in vitro-maturated porcine oocytes after intracytoplasmic sperm injection (ICSI), we examined the combined effect of U0126, a specific inhibitor of mitogen-activated protein kinase kinases, and an electrical pulse on pronuclear formation and developmental competence. Two approaches were tested: (i) 6-h treatment of ICSI oocytes with U0126 applied at different intervals (0, 2, 3 or 4 h) after the electrical pulse and (ii) treatment of ICSI oocytes with U0126 applied 4 h after the electrical pulse over an additional 4, 6 or 8 h. Another protein kinase inhibitor, 6-dimethylaminopurine, was used as a chemical activator in control experiments. The highest rates of diploid embryo formation, normal fertilization and blastocyst formation were observed after 6 h of exposure to U0126 starting 4 h after the electrical pulse. Therefore, U0126 can be used as an activating agent for porcine oocytes fertilized by ICSI.

RNAi screening identifies TAK1 as a potential target for the enhanced efficacy of topoisomerase inhibitors.

In an effort to develop strategies that improve the efficacy of existing anticancer agents, we have conducted a siRNA-based RNAi screen to identify genes that, when targeted by siRNA, improve the activity of the topoisomerase I (Top1) poison camptothecin (CPT). Screening was conducted using a set of siRNAs corresponding to over 400 apoptosisrelated genes in MDA-MB-231 breast cancer cells. During the course of these studies, we identified the silencing of MAP3K7 as a significant enhancer of CPT activity. Follow-up analysis of caspase activity and caspase-dependent phosphorylation of histone H2AX demonstrated that the silencing of MAP3K7 enhanced CPT-associated apoptosis. Silencing MAP3K7 also sensitized cells to additional compounds, including CPT clinical analogs. This activity was not restricted to MDA-MB-231 cells, as the silencing of MAP3K7 also sensitized the breast cancer cell line MDA-MB-468 and HCT-116 colon cancer cells. However, MAP3K7 silencing did not affect compound activity in the comparatively normal mammary epithelial cell line MCF10A, as well as some additional tumorigenic lines. MAP3K7 encodes the TAK1 kinase, an enzyme that is central to the regulation of many processes associated with the growth of cancer cells (e.g. NF- κB, JNK, and p38 signaling). An analysis of TAK1 signaling pathway members revealed that the silencing of TAB2 also sensitizes MDA-MB-231 and HCT-116 cells towards CPT. These findings may offer avenues towards lowering the effective doses of Top1 inhibitors in cancer cells and, in doing so, broaden their application.

Protection of red snapper (Lutjanus sanguineus) against Vibrio alginolyticus with a DNA vaccine containing flagellin flaA gene.

AIMS: The main aims of this study were to construct a DNA vaccine containing flagellin flaA gene from Vibrio alginolyticus strain HY9901 and to explore the potential application of pcDNA-flaA as a DNA vaccine candidate for red snapper (Lutjanus sanguineus). METHODS AND RESULTS: Plasmid DNA encoding flagellin flaA gene (designated as pcDNA-flaA) was used as a DNA vaccine to immunize red snapper. The distribution, expression and immunoprotection of the DNA vaccine were analysed in tissues of the red snapper by PCR, RT-PCR and challenge test. PCR results indicated that pcDNA-flaA distributed in liver, spleen, kidney, gill and injection site muscle at 7-28 days after vaccination. RT-PCR results indicated that the flaA gene was expressed in all above tissues of vaccinated fish at 7-28 days after vaccination. In addition, fish receiving the DNA vaccine developed a protective response to live V. alginolyticus challenge 28 days post inoculation, the relative per cent survival (RPS) was 88%. CONCLUSIONS: This study showed that injection of pcDNA-flaA induced an efficient, systemic and antigen-specific immune response in red snapper, which makes it an effective vaccine candidate against V. alginolyticus infection. SIGNIFICANCE AND IMPACT OF THE STUDY: The finding that red snapper does adequately respond to pcDNA-flaA intramuscular injection makes pcDNA-flaA a promising candidate for DNA vaccine treatment. Furthermore, the availability of red snapper for foreign gene expression represents a useful model to develop effective prophylactic strategies and opens new perspectives for the treatment of bacterial pathogens of marine cultured fish.

Identifying LRRC16B as an oncofetal gene with transforming enhancing capability using a combined bioinformatics and experimental approach.

Oncofetal genes are expressed in embryos or fetuses, are downregulated or undetectable in adult tissues, and then re-expressed in tumors. Known oncofetal genes, such as AFP, GCB, FGF18, IMP-1 and SOX1, often have important clinical applications or pivotal biological functions. To find new oncofetal-like genes, we used the public information of expressed sequence tags to systematically analyze gene expression patterns and identified a novel oncofetal-like gene, LRRC16B. It increased the proliferation, anchorage-independent growth and tumorigenesis of transformed cells in xenografts, possibly through its effects on cyclin B1 protein levels. These findings exemplify the feasibility of using bioinformatics to find new oncofetal-like genes and suggest that more genes with important functional roles will be uncovered in the candidate gene list.

Immunoproteomic analysis and identification of novel immunogenic proteins from Vibrio harveyi.

AIMS: The main aim of this study was to screen novel immunogenic proteins of Vibrio harveyi, which could be vaccine candidates. METHODS AND RESULTS: Whole-cell proteins of V. harveyi, strain Li01 and Huang01, were first separated by isoelectric focusing, followed by 2D-PAGE, respectively. Immunogenic proteins were identified by Western blotting, using Epinephelus coioides antisera against V. harveyi strain Li01. Western blot analyses revealed 16 shared immunogenic protein spots in both strains. All of the immunogenic proteins were successfully identified and corresponded to 15 proteins. None of these proteins have been previously reported as immunogenic for V. harveyi. Of the 15 proteins, 11 are specific immunoreactive proteins and four are nonspecific immunoreactive proteins. Furthermore, outer membrane protein N (spot 2) and oligopeptide ATP-binding cassette (ABC) transporter (spot 3) were used as immunogens to immunize E. coioides for investigation of their protective abilities and activities. The E. coioides immunized with OmpN has abilities to fight against infections caused by V. harveyi Li01 and Huang01. However, vaccination with oligopeptide ABC transporter induces low protective immune response in fish. CONCLUSIONS: Eleven novel specific antigens were found, and OmpN could potentially be used as vaccine candidate for the development of novel vaccine against V. harveyi. SIGNIFICANCE AND IMPACT OF THE STUDY: These data show that immunoproteomics methods can be successfully applied in identifying immunogenic proteins of V. harveyi, which helps to search for the protective antigens in future.

Fertility in single-ovulating and superovulated dairy heifers after insemination with low dose sex-sorted sperm.

The present study was performed to test fertility in single-ovulating and superovulated dairy heifers after insemination with low dose sex-sorted sperm under field conditions. Some parameters, including the dosage, deposition site and timing, were assessed with the pregnancy rates after artificial insemination (AI). Moreover, the use of oestrus synchronization in combination with sorted sperm was evaluated. Besides that, we also improved the embryo production efficiency in superovulated dairy heifers by optimizing the timing of inseminations and repartitioning the sexed sperm dosage among multiple inseminations. The conception rate (52.8%) in heifers after low dose (2 × 10(6) ) insemination with sorted sperm deep into the uterine horn did not differ (p > 0.05) from that (59.6%) of conventional AI (1 × 10(7) non-sorted sperm) and that of deep insemination with low dose non-sorted sperm (57.7%). There was also no difference (p > 0.05) between conception rates after single (51.7%) and double (53.8%) deep insemination with sorted semen. Heifers inseminated with sorted sperm at synchronous oestrus had a lower pregnancy rate (48.1%) than heifers at spontaneous oestrus (53.6%), but this did not reach statistical difference (p > 0.05). The average number of transferable embryos collected in vivo from heifers inseminated with sorted sperm (4.81 ± 2.04) did not differ (p > 0.05) from that obtained from heifers after insemination with non-sorted sperm (5.36 ± 2.74). Thus, we concluded that the pregnancy rate after deep intra-uterine insemination with low dose sorted sperm was similar to that of non-sorted sperm, which was either also deposited at a low dose deep intra-uterine or into the uterine body. Synchronization of oestrus can be beneficial in combination with sorted sperm to optimize the organization and management of dairy herds. The results from superovulated heifers demonstrated that our insemination regime can be used to obtain a comparable embryo production efficiency with sorted sperm than with non-sorted sperm.

Loop-mediated isothermal amplification method for rapid detection of Vibrio alginolyticus, the causative agent of vibriosis in mariculture fish.

AIMS: The purpose of this study was to develop a loop-mediated isothermal amplification (LAMP) method for the rapid, sensitive and simple detection of Vibrio alginolyticus in mariculture fish. METHODS AND RESULTS: LAMP primers were designed by targeting the gyrB gene. With Bst DNA polymerase, the target DNA can be clearly amplified for 60 min at 64 degrees C in a simple water bath. The detection sensitivity of the LAMP assay for the detection of V. alginolyticus is about 3.7 x 10(2) CFU ml(-1) (3.7 CFU per reaction). LAMP products could be judged with agar gel or naked eye after the addition of SYBR Green I. There were no cross-reactions with other bacterial strains indicating a high specificity of the LAMP. The LAMP method was applied to detect V. alginolyticus-infected fish tissues effectively. CONCLUSIONS: The LAMP established in this study is a simple, sensitive, specific, inexpensive and rapid protocol for the detection of V. alginolyticus. SIGNIFICANCE AND IMPACT OF THE STUDY: This LAMP method provides an important diagnostic tool for the detection of V. alginolyticus infection both in the laboratory and field.

Macrophage infiltrates with high levels of Toll-like receptor 4 expression in white adipose tissues of male Chinese.

BACKGROUND AND AIMS: Macrophages and Toll-like receptor 4 (TLR4) are involved in the inflammatory process of adipogenesis. This study aimed to characterize macrophage infiltrates and the associated TLR4 expression in different locations of white adipose tissues (WAT) of male Chinese and determine their correlations to adipocyte hypertrophy and hyperplasia. METHODS AND RESULTS: A total of 30 men, who were lean, overweight or with type 2 diabetes (T2D), were recruited. Their abdominal omental WAT (oWAT) and subcutaneous WAT (scWAT) were obtained. The contents of macrophages in oWAT and scWAT were quantified using anti-CD68 staining. The levels of TLR4 expression were analyzed by western blot assays and the adipocyte size was quantified, followed by linear regression analysis. Significantly higher numbers of macrophages (24.4+/-3.2 vs 6.1+/-2.9, p<0.001), associated with higher levels of TLR4 expression (0.59+/-0.19 vs 0.20+/-0.03, p<0.001), were observed in oWAT, as compared with that in scWAT. Furthermore, the levels of macrophage infiltrates and TLR4 expression in oWAT of subjects who were overweight or/and have T2D were significantly higher than that in the lean group. The average adipocyte diameters and cross-sectional areas in oWAT of subjects who were overweight were significantly greater than those in the lean group (p=0.003 and p=0.04, respectively). Importantly, the numbers of macrophage infiltrates were positively correlated to the levels of TLR4 expression, the sizes of adipocytes, the levels of body mass index and C-reactive protein, respectively. CONCLUSION: Our data suggest that macrophage-related TLR4 expression and inflammation contribute to the development of adipocyte hypertrophy in male Chinese.

Single-nucleotide polymorphism identification in the caprine myostatin gene.

Polymerase chain reaction (PCR) products of MSTN gene amplified from 35 goats representing 17 Chinese indigenous goat breeds and five imported goat breeds were sequenced to identify the single-nucleotide polymorphisms (SNPs) of a 379-bp fragment including part of intron 2 and exon 3 of MSTN gene. A total of eight SNPs (A1980G, G1981C, A1982G, G1984T, A2121G, T2124C, G2174A and A2246G) were identified among the sequenced goats. The SNPs found are all located in intron 2 except for A2246G, which was a synonymous mutation in exon 3. Four haplotypes were sorted from these eight SNPs, of which, haplotype I (AGAGATGA) and haplotype II (GCGTGTAA) are the two main haplotypes with the frequency of 77.8% and 14.8% respectively. The SNPs found at positions 1980, 1981, 1982, 1984 and 2121 might be linked to inheritance completely.