CN1227964C

CN1227964C - Breeding method for pure filement of laver

Info

Publication number: CN1227964C
Application number: CN 200410024354
Authority: CN
Inventors: 汤晓荣; 姜红霞
Original assignee: Ocean University of China
Current assignee: Ocean University of China
Priority date: 2004-06-15
Filing date: 2004-06-15
Publication date: 2005-11-23
Anticipated expiration: 2024-06-15
Also published as: CN1582642A

Abstract

The present invention relates to a method for producing seeds of pure filaments of laver. The present invention is characterized in that firstly, a thallus individual is selected out of a wild or culture porphyra haitanensis group, and the sex is determined; the selected alga is scrubbed by absorbent cotton balls in sea water; the scrubbed alga is pasted on specimen paper or copy paper, and is ventilated and dried in shade; an alga piece of 1 to 2mm is cut from the position near the middle base part of the alga by a scalpel, the alga piece is an alga without sundries by the inspection and the determining of imcroscope, and all cells are nutrient cells; the front piece which is cleaned by ultrasonic is cultured under the conditions of the temperature of 19 DEG C to 27 DEG C, the optical intensity of 2500 to 6000 lux, and the light irradiation time of 12L: 12D; red filament groups are grown after 1 to 2 months; the single alga group is separated to be cultured and enlarged. The present invention adopts unisexual individuals and nutrient cells, and has no introduction of foreign genetic materials in the whole seed production process. The obtained filament has heredity unicity, and has pollution-free ratio more than 95 %.

Description

A kind of producing method for seed of Porphyra haitanensis filamentous pure lines

Technical field

The present invention relates to a kind of producing method for seed of Porphyra haitanensis filamentous pure lines.

Background technology

Porphyra haitanensis is China's southern important laver cultivation kind, in Zhejiang, Fujian and Guangdong Coastal have extensive cultivation.The historical existing many decades of the artificial cultivation of Porphyra haitanensis, the cultivated species algae generally is with wild frond, promptly obtains to be used for the thallus seed of marine cultivation from the direct fruit picking spore inoculating of wild algae shell conchocelis.In recent years, also there is a few studies person in the laboratory, to obtain filamentous and is used to inoculate shell acquisition cultivation thallus seed, but this filamentous adopts male laver of a strain and the female laver of a strain to organize co-incubation and make it to hybridize and obtain, thereby the carpospore filamentous that produces is a heterozygosis; And in the thallus cultivated population heredity that obtains with such filamentous inoculation shell also be mix with uncertain, so thallus offspring's proterties instability directly causes laver output and the quality instability gathered in the crops, influences the producer's economic benefit.

Summary of the invention

The object of the invention provides a kind of producing method for seed that obtains Porphyra haitanensis filamentous pure lines, and it can overcome

The above-mentioned shortcoming of prior art.

A kind of producing method for seed of Porphyra haitanensis filamentous pure lines is characterized in that selecting the thallus individuality earlier from the Porphyra haitanensis colony of wild or cultivation, and definite sex; Selected frond is scrubbed with rayon balls in seawater; The frond of washing is attached on sample paper or the copy paper, and the frond that dries in the shade ventilates; The algae sheet is downcut in position with scalpel base portion in close frond, determines that at test under microscope the assorted algae of nothing and whole cell all are trophozytes; The algae sheet is cleaned with ultrasonic, and last microscopy determines that the two sides of tissue all do not have assorted algae or protozoa and adhere to; To handle clean a slice algae sheet at 19～27 ℃, light intensity 2500～6000lux cultivates under the condition of 12 hours every days of light application time; To there be red filamentous algae to fall to growing through 1-2 month time; Single algae is fallen to telling to cultivate amplification.

Advantage of the present invention: 1, because Porphyra haitanensis is a gonochorism, so the male and female trophozyte is distributed on the different individualities.What the present invention selected for use is the unisexuality individuality, and what choose is trophozyte, and whole production of hybrid seeds process does not have the introducing of foreign heredity substance; 2, used starting material is very little, and falls behind and separate obtaining the filamentous algae, single algae is fallen cultivate separately, can guarantee that the genetic origin of the filamentous that obtains is single.The homogeneous proterties that obtains the synchronism of thallophytic growth course and thallus offspring by filamentous has also shown the hereditary unicity of the filamentous that obtains.3, adopt ultrasonic to remove the assorted algae on surface, pollution-free rate reaches more than 95%.

Embodiment

The inventor takes back the laboratory after the female and male thallus individuality of the Porphyra haitanensis that picked up from Liaanjiang county, Fujian cultivated population in autumn in 2002 is dried in the shade, with-20 ℃ of freezing preservations behind the plastic sack sealed packaging.The frond that selection spring in 2003 is individual greatly, the frond color and luster is good, maturation is poor is in 20 ℃ of filtering seas, low light (being lower than 500lux) recovered after 1 day, select and have the purpose proterties individuality of (elongated, color and luster is shiny black, maturation is late) respectively, determine it is female or male at microscopically as individuality; In filtering sea, do preliminary cleaning to remove the main aufwuch in frond surface, generally need 2～3 times with tweezers and rayon balls; The frond that to choose is made sample on sample paper (if no sample paper, the white copy paper replacement of available similar size), and is placed on shady and cool ventilation place to the surface of frond and does not have the visible globule, and at this moment the frond cell still keeps normal vigor; Downcut vegetative tissue fritter (size is as far as possible little, and general 1～2mm's is square) with scalpel in the position near base portion in the frond then, it all is trophozyte that microscopy is determined; The algae sheet that obtains is placed in the small beaker of the centrifuge tube of 10ml or 50ml, adds the seawater (for 1/2～2/3 height of container) that boils the back cooling, with the mix removing of algae of ultrasonic crusher; In order to obtain higher success rate, can get 3～4 algae sheets at the different parts of each strain, be placed on simultaneously in the small beaker of a 50ml, add the seawater that boils the back cooling of 20～30ml; That the inventor uses is the ultrasonic crusher VC750 of U.S. SONICS-MAREIALS company, and using amplitude is 35%, handles 10 seconds totally 3 times at every turn, all changes water at every turn and cleans.After handling, the two sides of microscopy algae sheet does not all have assorted algae pollution and protozoa is promptly achieved the goal, if still have assorted algae, can repeat above ultrasonic treatment step, but in order to guarantee not damage the frond cell, should reduce to some extent each time that continues.Then, the triangular flask that every algae sheet is placed on 100ml is cultivated, and temperature is 20 ℃, and light intensity 2000lx, changed and once cultivates seawater at 12 hours every days of light application time in 2～3 weeks.After one month, all there is the filamentous algae to fall to growing on the bottle end and the algae sheet, single algae fallen to choosing in the triangular flask of a 250ml cultivate separately, can grow the filamentous of a great deal of after the several months, be purpose product filamentous pure lines.

Generally less by the thread scale of construction that male tissue obtains at first with method of the present invention, the time of separation can suitably postpone; And generally more by the thread scale of construction of female sex organization acquisition, fall for the ease of obtaining single algae, macroscopic algae falls and will separate as early as possible.That the developing synchrotron of the filamentous that obtains like this by further evidence is better than is wild (being heterozygosis) carpospore filamentous, this high efficiency seedling cultivating for artificial cultivation and production provides may.

Claims

1, a kind of producing method for seed of Porphyra haitanensis filamentous pure lines is characterized in that selecting the thallus individuality earlier from the Porphyra haitanensis colony of wild or cultivation, and definite sex; Selected frond is scrubbed with rayon balls in seawater; The frond of washing is attached on sample paper or the copy paper, and the frond that dries in the shade ventilates; The algae sheet is downcut in position with scalpel base portion in close frond, determines that at test under microscope the assorted algae of nothing and whole cell all are trophozytes; The algae sheet is cleaned with ultrasonic, and last microscopy determines that the two sides of tissue all do not have assorted algae or protozoa and adhere to; To handle clean a slice algae sheet at 19～27 ℃, light intensity 2500～6000lux cultivates under the condition of 12 hours every days of light application time; To there be red filamentous algae to fall to growing through 1-2 month time; Single algae is fallen to telling to cultivate amplification.