不同载体冷冻小鼠胚胎的效果比较

为了比较冷冻叶片(Cryoleaf)、冷冻帽(Cryotop)、自制半麦管(hemi-straw)三种冷冻载体的冻存效果,试验采用玻璃化冷冻方法冻存昆明小鼠8-cell胚胎。结果表明,Cryoleaf组、Cryotop组和hemi-straw组胚胎复苏率分别为92.3%、89.2%和86.2%,三组比较差异不显著(P0.05)。Cryoleaf组、Cryotop组及hemi-straw组胚胎的囊胚形成率分别是95.0%、91.4%和80.4%,三组比较差异显著(P0.05);新鲜对照组囊胚形成率为98.5%,与Cryoleaf组、Cryotop组分别比较差异不显著(P0.05),与自制hemi-straw组比较差异显著(P0.05)。结论:采用玻璃化冷冻方法冻存小鼠8-cell期胚胎,使用Cryotop和Cryoleaf作为冷冻载体,可以获得更高的复苏率和囊胚率,优于自制hemi-straw,可以很好地用来冻存人类的胚胎。     

猪卵母细胞在GV期与MⅡ期的冷冻保存效果比较研究

为比较猪卵母细胞在GV期与MⅡ期的冷冻保存效果,试验在这两个成熟阶段对其进行玻璃化冷冻,GV期卵母细胞解冻后培养至成熟,MⅡ期卵母细胞解冻后恢复2 h,然后采用免疫荧光标记、Western blotting和链霉蛋白酶溶解方法分别检测它们的皮质颗粒分布、CD9蛋白表达水平和透明带消化时间上的差异。结果表明,GV期卵母细胞在解冻后2 h的存活率显著低于MⅡ期卵母细胞（P<0.05）,但极体排出率与对照卵母细胞无明显差异（P>0.05）;在冷冻MⅡ期卵母细胞中,皮质颗粒的皮质区分布比例和CD9的蛋白表达水平显著下降（P<0.05）,但冷冻GV期卵母细胞经体外成熟后则无明显变化（P>0.05）;冷冻GV期与MⅡ期卵母细胞均不会影响透明带的消化时间（P>0.05）。由此可见,猪卵母细胞在GV期的冷冻存活率虽然较MⅡ期低,但其体外成熟后极体排出率、皮质颗粒分布和CD9蛋白表达水平均未受到冷冻的影响。     

乌腺金丝桃茎尖离体培养基激素配比筛选

为寻求乌腺金丝桃的高效人工繁殖方法,以乌腺金丝桃茎尖作为外植体,筛选初代诱导、继代增殖、生根培养3个阶段的最佳培养基激素配方。结果得出,把经过消毒后的乌腺金丝桃茎尖切割成0.3 mm、周围带2~3个叶原基,转入MS+6-BA 3.0 mg/L+NAA 0.1 mg/L+LH 500 mg/L培养基,分化率达到100%,能诱导出大量的健壮不定芽;然后将这些不定芽切割成1 cm小段转入MS+6-BA 2.0 mg/L+IBA 0.2 mg/L+GA_3 1.0 mg/L培养基,增殖效果最好;经过2~3次的继代增殖后,将增殖的大量苗木切割成2 cm小段转入到1/2 MS+IAA 0.2 mg/L生根培养基中,生根效果最佳,平均根数达到4.6,且根系粗壮,有利于今后出瓶移栽。     

树莓茎尖玻璃化法超低温保存及植株再生研究

为了建立适于树莓茎尖的超低温保存技术程序,试验以树莓茎尖为试材,对树莓玻璃化法超低温保存进行了研究。结果表明适于树莓的超低温保存技术流程是先低温驯化21d,在含0.8mol/L蔗糖的固体培养基预培养4d,再经玻璃化液(PVS2)处理120min后浸入液氮,解冻后茎尖存活率达75%。     

Cryopreservation of Queen Honeybee （Apis mellifera carnica） Born Worker Eggs by Vitrification

Many species of insect egg can be targeted individually or （and） collectively for cryopreservation by vitrification. However, there has been no report on cryopreservation of honeybee eggs by vitrification. In an attempt to define a preliminary procedure of cryopreservation of honeybee eggs by vitrification, queen honeybee born worker eggs （worker eggs） were stored through vitrification in liquid nitrogen up to 1 h, and then post-vitrification survival of the vitrified worker eggs in vitro and their hatching rates during maturation in vivo were observed using microscopic and close visual inspections. The procedure of cryopreservation by vitrification included dechorionation with sodium hypochlorite and permeabilization with isopropyl alcohol; equilibration by addition of loading solution （i.e., 25% vitrification storage solution） and dehydration by gradual replacement of loading solution with vitrification storage solution; cooling in liquid nitrogen vapor right before droplet vitrification in liquid nitrogen; and recovery in liquid nitrogen vapor right after storage in liquid nitrogen, thawing at temperature of thawing medium （5% sucrose in TC 100-insect medium） and rehydration by gradual replacement of vitrification storage solution with rehydration solution （5% fetal bovine serum in TC 100-insect medium）. It was found that among the worker eggs experiencing cyropreservation by vitrification, 1.25% of them were successfully passed through the four life stages, viz., egg, larva, pupa, and adult. In summary, it can be inferred that although a majority of worker eggs were dead after cyropreservation by vitrification, a few of them were developed into larvae, pupae, and finally emerged as adults.     

动物胚胎冷冻保存技术的研究进展

分析了动物胚胎冷冻保存的机理,综述了几种常见的动物胚胎冷冻保存方法的研究进展,并对动物胚胎玻璃化冷冻保存技术的发展前景进行了展望。     

密花石斛花粉的保存方法研究

研究了密花石斛花粉不同保存方法下的花粉寿命,并对密花石斛花粉的超低温保存技术程序进行了研究。 结果表明:1)密花石斛花粉采用室温（(27±2)℃）、冷藏（4℃）、冷冻（-20℃）保存,花粉寿命分别为5、23、17 d,超低温保存(-196℃)的花粉120 d后仍可检测到花粉萌发,且萌发率与新鲜花粉相同,超低温保存方法至少可以保存密花石斛花粉4个月;2)密花石斛花粉超低温保存的程序为新鲜花粉（含水量43.26%）直接投入液氮（LN）保存,需用时采用室温、自来水或水浴化冻即可。      

甘薯茎尖超低温保存技术研究

本试验采用包埋玻璃化法超低温冷冻技术保存徐薯18号的茎尖,结果表明,当剥取茎尖大小为1.0~1.5 mm,玻璃化处理时间为90 min时,茎尖存活率达85%以上。     

影响猪卵母细胞玻璃化冷冻保存效果的因素

对卵母细胞进行冷冻保存,可以使卵源不再受到时间和空间限制,并能为体外受精、核移植和转基因等胚胎工程技术的研究提供充足的原材料.猪卵母细胞中富含脂滴,且对低温十分敏感,因此,猪卵母细胞冷冻保存的研究进展相对较慢.玻璃化冷冻是有效的卵母细胞冷冻方法之一.综述了卵母细胞发育阶段、冷冻栽体和冷冻保护剂等因素对猪卵母细胞玻璃化冷冻保存效果的影响.     

青蒿组织培养中克服玻璃化现象研究

玻璃化现象是青蒿组织培养过程中的首要障碍。用青蒿顶芽为外植体,以MS为基本培养基,采用不同浓度的6-BA、琼脂、蔗糖和活性炭正交实验对解决青蒿组织培养过程中的玻璃化问题进行了研究。结果表明:在MS培养基添加4.0%的蔗糖、0.75%琼脂和0.08%的活性碳,附加0.10 mg/L的细胞分裂素6-BA,可得到健壮的青蒿组培苗。